Methods for production of the oxidized glutathione composite with cis-diamminedichloroplatinum and pharmaceutical compositions based thereof regulating metabolism, proliferation, differentiation and apoptotic mechanism for normal and transformed cells

ABSTRACT

The present invention relates to a composite for the treatment of a variety of medical conditions, the composite comprising an oxidized glutathione-based compound, which has a disulfide bond, and a metal material, in particular where the metal is either platinum or palladium. The oxidized glutathione-based compound and metal material can be present in a ratio of 3000 to 1 and preferably 1000 to 1. The oxidized glutathione-based compound can be oxidized glutathione itself or salts or derivatives. A feature of the invention is that the composite has a more stabilized disulfide bond than the oxidized glutathione-based compound itself. Methods for preparing the composite are provided, such methods being beneficial in that the composite is provided in high yields and at high purity. Methods for treating various medical conditions with the composites of the present invention are also disclosed.

RELATED APPLICATIONS

[0001] This application is a continuation of U.S. application Ser. No.09/241,232, filed Feb. 1, 1999, now pending, which is acontinuation-in-part of U.S. application Ser. No. 09/237,801, filed Jan.27, 1999, now abandoned.

FIELD OF THE INVENTION

[0002] The present invention relates to medicine and, more particularly,to pharmacology, i.e., to methods (or producing medicinal agents basedon a composite comprising an oxidized glutathione composite and aplatinum material, in particular cis-platin, that are intended to beused for preventing and treating various pathologic syndromes anddiseases by way of differentiated influence on processes of metabolism,proliferation, differentiation and apoptosis of normal and transformedcells.

BACKGROUND OF THE INVENTION

[0003] Certain issues in modern pharmacological industry involvecounteracting two medical-biological problems:

[0004] forming a resistance (tolerance and pharmacological efficacydecrease) to medicinal agents including cases due to activation of theMDR-genes system; and

[0005] forming of undesirable resorptive effects manifested, first ofall, by alteration of the immunocompetent cell system and hemopoiesis;cardio-, hepato-, nephro- and neurotoxicity.

[0006] A typical cause for these problems is a wide administration ofchemotherapeutic agents that can be quite effective, according to theirphysical-chemical properties, but can also be foreign to an organism attheir very nature. Even genetic-engineering medicines, in spite of humangenes (DNA) application as a matrix for multiplication, usesingle-celled organisms (Escherichia coli, yeast cells) that bring intheir own, and therefore, xenobiotic contribution into obtained drugs.

[0007] Theoretical and practical medicinal research are now payinggreater attention to natural key metabolites, i.e., key factors (lowmolecular biochemical substances) that are naturally determined totrigger chain reactions for endogenous production and modification ofmany biologically active products for physiologically important andadequate processes. In some cases these biochemical substances functionas “biochemical gyroscopes” assigned to restoring the balance of alteredequilibrium of basic metabolic processes, for example, anabolism andcatabolism or proliferation and differentiation. Alteration of thebalance of these vital mechanisms can lead to cell destruction(cytolytic syndrome) or their transformation into malignant ones, i.e.,cancer. As a rule, the key regulatory metabolites, i.e., “cellularhormones”, are peptide origin factors (usually not larger than 3-20amino acids). Obtaining synthetic analogues (biochemical substances)and, therefore, drugs with predetermined properties is a desired goalbecause these drugs are optimal as metabolic therapy instruments and, infact, are not foreign to the organism.

[0008] Ideal peptide structures that can function as these syntheticanalogues include sulphur-containing peptides and derivatives thereof,due to the presence of a thiol group. In particular, biological effectsof the tripeptide “reduced glutathione” (γ-glutamyl-cysteinyl-glycine;hereinafter—GSH) are known to be researched extensively. The glutathionetripeptide dimer, oxidized glutathione (γ-glutamyl-cysteinyl-glycine;hereinafter—GSSG), where two molecules of the tripeptide with theaforementioned formula are linked via a covalent bond between cysteineresidues, is also well known.

[0009] Administration of the exogenous synthetic GSSG analogue is knownto induce cytokine and hemopoietic factor synthesis during in vitroexperiments, and to provide the setting of the cytokine profile tonormal values in conditions of cyclophosphamide and radiationimmunodepression models (in vivo experiments) along with the immunityand hemopoietic system restoration (International application WO97/21444,MKI A61 κ 38/02, published Jun. 19, 1997).

[0010] In their turn, the exogenous GSSG drug forms applied to severeimmunodeficient conditions and suppressed bone marrow hemopoiesiscontaining chemical examples relating to clinical Examples in theInternational Application on AIDS patients, patients with oncopathology,aplastic anemia,and other conditions at treatment courses of differentdurations, can provide a curative effect in restoring the organismimmune status (including antitumor immunity indices), immunogenesis andhemopoietic functional activity (International application WO 97/21444,MKI A61 κ 38/02, published Jun. 19, 1997).

[0011] Previously, the thiol class of biologically active substance hadapplications aimed to provide GSH at increased levels, i.e., to obtainantioxidant effects. In addition, pharmacological activity was observed,namely, for the pro-oxidant effect gaining and forming of a newintracellular redox-balance by introduction of GSSG that possessespro-oxidant potential into an organism. This is the only known case oftriggered redox-sensitive mechanisms for immunologically significantgenes, activation of the cellular thiol metabolism, and therefore,beneficial pharmacological properties were provided including systemiccell-protective effects and immunity state regulation depending oninitial cell status: immunodeficiency, i.e., hyporeactivity,immunoautoaggression, i.e., hyperreactivity.

[0012] International Application WO 97/21444,MKI A61 κ 38/02, publishedJun. 19, 1997, is directed to gain a set of technical andpharmaceutically acceptable solutions effective for the prevention ofthe GSSG reduction into GSH and, thus, for extending the lifetime ofGSSG as the oxidized form in biological media. Attainment of thebiological-pharmacological effects of the glutathione oxidized form isproven by the biomedical investigation results obtained in the course ofthe complex and extensive preclinical and clinical studies program onsynthetic GSSG analog effectiveness. A strategy for extending thelifetime of oxidized GSSG includes providing: salts thereof, compositedrugs including GSSG combinations with substances that prolong orenhance the effect of GSSG or salts thereof, or derivatives as a newcomposite, i.e., a mixture of oxidized GSSG and other materials. Thepharmaceutically acceptable GSSG derivative in the form of the saltsthereof, or combinations with extenders of the GSSG existence in theoxidized form, or GSSG combinations with enhancers/modifiers, i.e., alltechnical solutions stabilizing in the varying degree the GSSG moleculedisulfide form were first demonstrated to be significantly moreeffective in inducing the cytokine and hemopoietic factor production innormal conditions and to a greater degree in pathologic ones.

[0013] GSSG derivative drug forms are characterized with a largerfraction of the GSSG stabilized in the disulfide form, with maximalpharmacokinetics in biological media. These forms manifested thefollowing features:

[0014] a) Inducting production of a wider range of cytokine andhemopoietic factors that can determine the presence of largelymodulating effects rather than only stimulating ones.

[0015] b) Reproduction of particular cytokine effects, for instance,IL-2.

[0016] The events developing in cells (tissues and, therefore, organs)after interaction with cytokines is well known. These events areconditioned by the universal cytokine influence on the mainsignal-transducing systems and, through the latter, on the cell genomedetermining regulating cytokine effects on proliferation,differentiation, and apoptosis.

[0017] Methods for obtaining the oxidized GSSG form from the reduced GSHprecursor are well known. The labile mercapto-SH-groups of cysteine inGSH can be oxidized with such soft oxidizers as air oxygen (R. Douson,D. Elliot, W. Elliot, K. Jones. Biochemist's manual, Moscow, “Mir”,1991; Tam J. P. et al., Int. J. Pept. Prot. Res., Vol. 29, p. 421-431,1987; Ahmed A. K. et al., J. Biol. Chem. 250, p. 8477-8482, 1975).However, the reaction rate is rather low in this case and desiredquantitative yields require very long periods of time (many days).Catalysis by heavy metals ions and, especially, by copper can be toxic,and can create significant problems for obtaining pure pharmaceuticalmedicines.

[0018] Another oxidation method involves more potent oxidizers such as,hydrogen peroxide, iodine, potassium ferrocyanide, etc. (Kamber B. etal., Helv. Chim. Acta., Vol. 63, p. 899-915, 1980; Hope D. B. et al., J.Biol. Chem., Vol. 237, p. 1563-1566, 1962). These reactions generallyproceed much faster (dozens of minutes to several hours). Adisadvantage, however, is a difficulty in controlling reactionconditions that can result in significant contamination of the productwith oxidation products, e.g., derivatives of the corresponding acids.It is sometimes necessary to add additional, sometimes ratherlabor-consuming purification procedures, that can sharply appreciate theprocess.

[0019] Yet another oxidation method involves the use of gaseoussubstances (nitrogen oxides), sulfoxides and other compounds asoxidizers. These oxidizers, however, can be toxic. (William A. Kato etal., Chem. Pharm. Bull., vol. 34 (2), p. 486-495, 1986; A. Meister etal., Ann. Rev. Biochem., p. 711-718, 1983.)

[0020] All these methods do not necessarily improve the quantitativeyield of the desired product compared to older methods, and at the sametime, can provide additional problems regarding toxicity and safety, asin case of nitrogen oxides, or more difficult accessibility of thereagents and their high cost.

[0021] Another previously known method involves the use of hydrogenperoxide as an oxidizer (Amber B. et al., Helv. Chim. Acta., Vol. 63, p.899-915, 1980). The process is performed in the water solution with pHabout 8.0-8.5 using the hydrogen peroxide equivalent at the roomtemperature. The reaction time is about 1 hour and the product yield is90%. The main impurities (up to 10%) are other oxidation products, whichcan be removed only by means of an expensive preparative HPLC separationthat can sharply increase the drug cost.

SUMMARY OF THE INVENTION

[0022] The present invention involves the creation of newpharmaceutically acceptable compounds with predetermined propertiesbased on GSSG, i.e., an oxidized glutathione-based compound having astabilized disulfide bond. The present invention also provides a newmethod for obtaining the oxidized glutathione as a composite with thestabilized disulfide bond during the product synthesis. The methodinvolves the production of the composite having the formula:bis-(γ-L-glutamyl)-L-cysteinyl-bis-glycine disodium salt with a platinummaterial such as cis-platin, i.e., cis-diamminedichloroplatinum,preferably in the mole ratio 3000:1, more preferably in a mole ratio of1000:1.

[0023] According to the invention the composite is characterized ashaving a stabilized disulfide bond. Upon introduction into biologicalmedia. consequently, a longer drug half-life time is provided in thebiological media in the disulfide form.

[0024] The general procedure of the present method for the compositeproduction involves using the reduced glutathione for the oxidationreaction as a hydrogen peroxide oxidizer combined with a platinummaterial, in particular, cis-diamminedichloroplatinum, as a catalyst.The method allows using lesser amounts of hydrogen peroxide (forexample, 0.9 of an equivalent), resulting in an elimination of thesuperoxidation products along with very high yield for GSSG (more than98% by the HPLC data). Thus, the product obtained has high purity anddoes not require additional purification.

[0025] One aspect of the present invention provides a compositecomprising an oxidized glutathione-based compound and a metal materialin a ratio between about 3000:1 to about 1:1. The metal materialcomprises a metal selected from the group consisting of platinum andpalladium.

[0026] Another aspect of the invention provides a method for stabilizinga disulfide bond of an oxidized glutathione-based compound. The methodcomprises interacting the oxidized gIutathione-based compound with ametal material. The metal material comprises a metal selected from thegroup consisting of platinum and palladium.

[0027] Another aspect of the present invention provides a method ofstimulating endogenous production of cytokines and hemopoietic factors.The method comprises the steps of introducing to a mammalian body, inneed of stimulation of cytokines or hemopoietic factors or both, aneffective amount of a composite. The composite comprises an oxidizedglutathione-based compound and a metal material in a ratio of betweenabout 3000:1 to about 1:1. The metal material comprises a metal selectedfrom the group consisting of platinum and palladium. The method iscarried out for a period of time to stimulate the endogenous productionto obtaining a therapeutic effect.

[0028] Another aspect of the present invention provides a method forenhancing and prolonging the ability of an oxidized glutathione-basedcompound to stimulate endogenous production of cytokine and hemopoieticfactors. The method involves the steps of interacting the oxidizedglutathione-based compound with a metal material in a ratio of betweenabout 3000:1 to about 1:1. The metal material comprises a metal selectedfrom the group consisting of platinum and palladium.

[0029] Another aspect of the present invention provides a method fortreating a subject having a disease. The method comprises administeringto the subject in need of such treatment a composite comprising anoxidized glutathione-based compound and a metal material in a ratio ofbetween about 3000:1 to about 1:1 in an amount effective to stimulateendogenous production of cytokines and/or hemopoietic factors or both toobtain the therapeutic effect. The metal compound has a metal selectedfrom the group consisting of platinum and palladium.

[0030] Other advantages, novel features, and objects of the inventionwill become apparent from the following detailed description of theinvention when considered in conjunction with the accompanying drawings,which are schematic and which are not intended to be drawn to scale. Inthe figures, each identical or nearly identical component that isillustrated in various figures is represented by a single numeral. Forpurposes of clarity, not every component is labeled in every figure, noris every component of each embodiment of the invention shown whereillustration is not necessary to allow those of ordinary skill in theart to understand the invention.

BRIEF DESCRIPTION OF THE DRAWINGS

[0031]FIG. 1 shows the structure ofbis-(γ-L-glutamyl)-L-cysteinyl-bis-glycine disodium salt withcis-diamminedichloroplatinum;

[0032]FIG. 2 shows the synthesis scheme of the composite of the oxidizedglutathione disodium salt with cis-diamminedichloroplatinum;

[0033]FIG. 3 shows the donor-acceptor bond among platinum atom and twoNH₃ groups due to the lone-pair electrons of nitrogen atoms;

[0034]FIG. 4 shows proposed mechanisms for the GSSG molecule disulfidebond stabilization due to the ligand exchange—NH₃-groups on disulfidebonds—and through forming of the donor-acceptor bond among platinum atomand two sulfur atoms due to lone-pair electrons of sulfur atoms;

[0035]FIG. 5 shows proposed mechanisms of the general GSSG moleculestabilization through the mechanism given at FIG. 4 as well as throughexchange of the NH₃ ligands on NH₂ groups of the glutathione (the NH₂group convergence and the GS fragments, correspondingly) forming new“biophysics” of the GSSG.Pt composite, with squares (□) denoting donorsites and circles (◯) denoting acceptor sites;

[0036]FIG. 6 shows the main sites (encircled) for the GSSG.Pt moleculechemical modification;

[0037]FIG. 7 shows the structure of bis-phenylalanyl-GSSG.Pt;

[0038]FIG. 8 shows the synthesis scheme forbis-(L-phenylalanyl-γ-L-glutamyl)-L-cysteinyl-bis-glycine withcis-diamminedichloroplatinum;

[0039]FIG. 9 shows the structure of lithium salt ofbis-(γ-L-glutamyl)-L-cysteinyl-bis-glycine;

[0040]FIG. 10 shows the stage of processing of the source reducedglutathione with hydrogen peroxide at pH=8 (stage of the synthesisscheme for the GSSG.Pt lithium salt);

[0041]FIG. 11 shows the extraction of the free hexapeptidebis-(γ-L-glutamyl)-L-cysteinyl-bis-glycine (III, stage of the synthesisscheme for the GSSG.Pt lithium salt);

[0042]FIG. 12 shows the transfer of the oxidized GSSG (III) form in thelithium salt of the GSSG.Pt;

[0043]FIG. 13 shows the DNA degradation character of the normal cells atthe control group (1), after treatment with: GSSG (2), GSSG.Pt (3)incubation time—48 hrs;

[0044]FIG. 14 shows the DNA degradation character of the HL-60 cells atthe control group (2), after treatment with: GSSG (1), GSSG.Pt (3)incubation time—48 hrs;

[0045]FIG. 15(a) shows the structure of S-thioethylamine•glutathionedisulfide;

[0046]FIG. 15(b) shows the structure of bis-[DL-6,8-thioeticacid]•glutathione disulfide;

[0047]FIG. 15(c) shows the structure of[β-alanyl-L-histidyl]•glutathione disulfide;

[0048]FIG. 15(d) shows the structure of[9-β-D-ribofuranosyladenyl]•glutathione disulfide;

[0049]FIG. 15(e) shows the structure ofbis-[L-2-amino-4-[methylthio]butanoic acid]. glutathione disulfide;

[0050]FIG. 16(a) shows the structure of bis-[methionyl]•glutathionedisulfide;

[0051]FIG. 16(b) shows the structure of bis-[aspartyl]•glutathionedisulfide;

[0052]FIG. 16(c) shows the structure of bis-[histidyl]•glutathionedisulfide;

[0053]FIG. 16(d) shows the structure ofbis-[3-iodine-tyrosyl]•glutathione disulfide;

[0054]FIG. 16(e) shows the structure of [γ-aminobutanoyl]•glutathionedisulfide;

[0055]FIG. 16(f) shows the structure ofbis-[γ-hydroxybutanoyl]•glutathione disulfide;

[0056]FIG. 16(g) shows the structure ofbis-[3,4-dihydroxyphenylalaninyl]•glutathione disulfide;

[0057]FIG. 17(a) shows the structure of bis-nicotinoyl-glutathionedisulfide (bis-[pyridine-3-carbonyl]•glutathione disulfide);

[0058]FIG. 17(b) shows the structure ofuridine-5′-monophosphatoyl•glutathione disulfide;

[0059]FIG. 17(c) shows the structure ofinosine-5′-monophosphatoyl•glutathione disulfide;

[0060]FIG. 17(d) shows the structure of folliculylsuccinyl•glutathionedisulfide;

[0061]FIG. 17(e) shows the structure ofglycerol-1,3-diphosphatyl•glutathione disulfide;

[0062]FIG. 18(a) shows the structure of tetra-dopamine•glutathionedisulfide;

[0063]FIG. 18(b) shows the structure of theophylline•glutathionedisulfide;

[0064]FIG. 19(a) shows the structure of bis-[carnosyl]-GSSG;

[0065]FIG. 20(a) shows the structure of a non-symmetric mixed disulfidecompound;

[0066]FIG. 20(b) shows the structure of a symmetric mixed disulfidecompound;

[0067]FIG. 20(c) shows the structure of a symmetric and doubly bridgedmixed disulfide compound;

[0068]FIG. 21(a) shows the structure of divanadate salts;

[0069]FIG. 21(b) shows the structure of dihydrofluoride salts;

[0070]FIG. 21(c) shows the structure of dilithium salts;

[0071]FIG. 21(d) shows the structure of dizinc salts;

[0072]FIG. 22 shows a scheme for modifying a GSSG.Pt composite havingphenylalanyl groups;

[0073]FIG. 23 shows a GSSG pharmokinetic curve for intravenousintroduction;

[0074]FIG. 24 shows a GSSG pharmokinetic curve for intravenousintroduction;

[0075]FIG. 25 shows a chart of immune system response to cancers anddiseases;

[0076]FIG. 26 shows a chart of drug activation of immune systems tofight cancers and diseases; and

[0077]FIG. 27 shows a chart of cytokines stimulated by drug-type.

DETAILED DESCRIPTION

[0078] The present invention discloses a number of oxidizedglutathione-based compounds having a stabilized disulfide bond and inparticular, a composite comprising the compound of this invention with ametal material. Methods for the preparation of a composite and for usingthe composite to treat a variety of diseases are also disclosed.

[0079] One aspect of the present invention provides a compositecomprising an oxidized glutathione-based compound and a metal material.Generally, a “composite” can refer to a mixture of different chemicalspecies. The “mixture” can be a physical mixture or a chemical mixture,i.e., having a chemical interaction involving either a chemical bond oran electrostatic interaction. In one embodiment, the mixture can beprepared by dissolving and/or suspending the different chemical speciesin a solution and precipitating out or filtering out a resulting solid.In another embodiment, the mixture can be a homogeneous solutioncomprising the different chemical species.

[0080] An “oxidized glutathione-based compound” refers to any compoundhaving a basic dimer structure where each unit of the dimer comprises aglutamic acid group or salt or derivative bonded to a cysteine group orsalt or derivative bonded to a glycine group or salt or derivative, andeach unit is correspondingly bonded to each other by the cysteine sulfuratoms to form a sulfur-Sulfur bond (disulfide bond). A “derivative” canbe prepared by reacting at least one reactive group of the oxidizedglutathione-based compound or precursor with another chemical species.An example of an oxidized glutathione-based compound is GSSG itself.Other examples are provided below.

[0081] In one embodiment, the oxidized glutathione-based compound hasthe general formula:

[0082] A, B, D, E, G and H can each be selected from the groupconsisting of an organic unit and salts of the organic unit. Preferably,the “organic unit” allows the glutathione-based compound to remainsoluble in biological media and in addition, the organic unit should notimpart toxicity to the oxidized glutathione-based compound in an applieddosage. It is understood that A, B, D, E, G and H can be the same ordifferent. Preferably, groups A—H can each include a unit selected fromthe group consisting of amine groups, carboxyl groups, and amides. Forexample, A—H can represent amino acids or derivatives bonded via anamide bond. Alternatively, any two of A—H can be linked to each other byat least one covalent bond. Thus, A—H can be part of a cyclic structure.

[0083] In one embodiment, the composite comprises a large excess of theoxidized glutathione-based compound relative to the metal material,preferably in a ratio of between about 3000:1 and 1:1, more preferablyin a ratio of between about 1000:1 and 1:1, more preferably in a ratioof between about 1000:1 and 10:1, even more preferably in a ratio ofbetween about 1000:1 and 100:1. In another embodiment, the compositecomprises equal amounts of the oxidized glutathione-based compound andthe metal material, i.e., a ratio of about 1:1.

[0084] In one embodiment, the metal material comprises a metal selectedfrom the group consisting of platinum and palladium. Preferably, themetal is platinum. Ideally, the metal material, in combination with theoxidized glutathione-based compound, renders the composite soluble inbiological media. Small portions of the metal material can be insoluble,as long as the insoluble portion does not result in any toxic orhazardous effects to the biological system. A platinum material can beselected from the group consisting of a platinum salt, a coordinationcompound and an organometallic compound. Preferably, the platinummaterial is a platinum coordination compound such as cis-platin(cis-Pt(NH₃)₂Cl₂ or cis-diamminedichloroplatinum—FIG. 3).

[0085] In one embodiment, the oxidized glutathione-based compound isoxidized glutathione itself (GSSG) and salts thereof, where both A and Eare —CO₂H, both B and D are —NH₂ and both G and H are —CO₂M, M being acounterion. The counterion can be a proton, an organic-based ion such astetralkyl-amononium, an alkaline metal, an alkaline earth metal, or atransition metal. It is understood that in aqueous media, any of A—H cancomprise an ionized group, e.g., A and E can be —CO₂ ⁻, and B and D canbe —NH₂ ⁺ and the ionized groups are neutralized by an appropriatecounterion.

[0086] In a preferred embodiment, the present invention relates to theproduction of a new biologically active compound, i.e., a compositecomprising an oxidized glutathione-based compound andcis-diamminedichloroplatinum. A convenient short-form notation will beused herein. For example, a composite comprising GSSG itself andcis-platin will be denoted as GSSG.Pt. Derivatives will be denoted bythe newly appended chemical group, e.g., bis-[histidyl]-GSSG. While notwishing to be bound by any theory, a proposed structural formula can befound in FIG. 1. (Gross-formula: C₂₀H₃₀N₆O₁₂Na₂S₂.[Pt(NH₃)₂Cl₂];molecular weight: 656.59 on C₂₀H₃₀N₆O₁₂Na₂S₂ with Pt content0.022-0.042%.)

[0087] The composite can be prepared by various methods. For example,the composite can result from the addition of a metal material toglutathione in the presence of an oxidant. Alternatively, the compositecan result by the addition of a metal material to the oxidizedglutathione-based compound.

[0088] Thus, another aspect of the invention provides a method forstabilizing a disulfide bond of an oxidized glutathione-based compound.“Stabilizing a disulfide bond” refers to a process for maintaining abond between two sulfur atoms and preventing facile reversion of theoxidized glutathione-based compound (e.g., GSSG) back to the reducedform (e.g., GSH). Alternatively, stabilizing the disulfide bond canresult in an increased lifetime of the oxidized glutathione-basedcompound. In the presence of reductants, the disulfide bond can cleaveresulting in formation of the reduced form of the glutathione-basedcompound, which is an undesired reaction. By maintaining theglutathione-based compound in an oxidized form for a greater amount oftime, the compound can be pharmaceutically effective for acorrespondingly longer period of time in biological media.

[0089] In one embodiment, the disulfide bond can be stabilized byinteracting the oxidized glutathione-based compound with a metalmaterial. As discussed previously, the metal material is preferably aplatinum or palladium material, such as cis-platin. In one embodiment,the platinum material is present in an amount of between about 0.0003equivalent to about 1 equivalent relevant to the oxidizedglutathione-based compound, preferably in a ratio of between about 0.001equivalent to about 1 equivalent relevant to the oxidizedglutathione-based compound.

[0090] In one embodiment, “interacting the oxidized glutathione-basedcompound with a metal material” comprises providing a glutathione-basedcompound and reacting this compound with an oxidant and a platinummaterial. A “glutathione-based compound” refers to any compound having astructure comprising a glutamic acid/salt/derivative bonded to acysteine/salt/derivative bonded to a glycine/salt/derivative. Examplesof glutathione-based compound include glutathione itself or anyderivative, where a derivative can be prepared by reacting a reactivegroup with another chemical species. The resulting product will be anoxidized glutathione-based compound having a stabilized disulfide bond.Thus, in this embodiment, a glutathione-based compound is in a reducedform, such as GSH, and the reaction with a oxidant involves oxidizingthe glutathione-based compound to produce a sulfur-sulfur bond. Theoxidant can be any species which can cleave a S—H bond of aglutathione-based compound to produce a hydrogen atom and a compoundhaving a sulfur-based radical which ultimately can react with anothersulfur-based radical to provide the sulfur-sulfur bond. Various oxidantsthat can perform this S—H bond cleavage are well known in the art. In apreferred embodiment, the oxidant is selected from the group consistingof oxygen and hydrogen peroxide.

[0091] In this method, reacting a glutathione-based compound with anoxidant and the platinum material comprises an oxidation reaction.Relative amounts of the reactants are preferably about 1 equivalent ofthe glutathione-based compound with less than about 1 equivalent of theoxidant such as hydrogen peroxide, and more preferably, with about 0.9equivalent of the hydrogen peroxide. In another embodiment, theoxidation reaction comprises reacting about I equivalent of theglutathione-based compound with between about 0.0003 equivalent andabout 1 equivalent of the platinum material, preferably between about0.001 equivalent and about 1 equivalent of platinum material, morepreferably between about 0.001 equivalent and about 0.1 equivalent, andeven more preferably between about 0.001 equivalent and 0.01 equivalent,in the presence of less than 1 equivalent of the oxidant. In anotherembodiment, about 1 equivalent of the glutathione-based compound isreacted with about 1 equivalent of the platinum material in the presenceof less than 1 equivalent of the oxidant.

[0092] In one embodiment, the method involves oxidizing theglutathione-based compound with about 0.9 equivalent of hydrogenperoxide and about 0.001 equivalent of cis-platin. One advantageousfeature of this method is an increased rate of oxidation of theglutathione-based compound. Another advantageous feature of this methodis that the yield of the resulting composite is increased to an amountgreater than about 98% and this increased yield is accompanied by anincreased purity. The purification of this composite is simplified to asignificant degree in that liquid chromatography can be performed toobtain a purity of the composite of greater than 99%, which complieswith pharmaceutical standards. Prior art methods have achieved a purityof only 75-93% of oxidized glutathione, depending on the method.

[0093] In a preferred embodiment, the composite is synthesized in onestep by oxidizing the reduced glutathione in the presence ofcis-diamminedichloroplatinum, which may function as an oxidationreaction catalyst. The reaction conditions can be regulated accuratelyby using less than 1 equivalent of hydrogen peroxide. Consequently,formation of superoxidation products can be reduced, resulting in a nearquantitative yield of the product. Thus, the one-step compositesynthesis provides significant technological simplification andproduction of the composite GSSG.Pt with the stabilized disulfide bond.

[0094] In a preferred embodiment, the reaction is performed in asolution involving reduced glutathione as a monosodium salt and addingabout 0.9 equivalent of the hydrogen peroxide and about 0.001 equivalentof cis-diamminedichloroplatinum at room temperature with stirring. Theoxidation reaction typically proceeds in about 1.5-2 hours. Control forthe completeness of the oxidation process can be conducted by an HPLCassay. The process is completed by the reaction solution lyophilicdrying to produce the composite consisting of oxidized glutathione andcis-diamminedichloroplatinum in a mole ratio of 1000:1 (confirmed byspectral analysis on platinum and sodium). The peptide constituent ofthe obtained composite according to the data of an amino acid assay, aNMR (¹H) spectrum, retention time by HPLC corresponds to GSSG. Theadmixtures content do not exceed 2%, and the product yield as a disodiumsalt is 96-98% calculating for the dry composite.

[0095] A composite synthesis scheme can be found on FIG. 2.

[0096] While not wishing to be bound by any theory, the increasedstabilization of the disulfide bond can be the result of an interactionof the sulfur atoms with the platinum material (see also FIG. 4).

[0097] In an interaction between the platinum material and the oxidizedglutathione (GSSG) molecule there is a possibility for ligand exchange,i.e., instead of the NH₃ groups, two sulphur atoms possessing two pairsof the lone-pair electrons can be involved in donor/acceptor bonds withthe platinum atom. One can also consider the possibility for theaddition in regard to the aforesaid stabilization of the disulfide bonddue to the convergence of the NH₂ groups of the oxidizedglutathione-based compound and stabilization of the general GSSGconformation (FIG. 5).

[0098] It is an advantageous feature of the present invention thatobtaining derivatives of GSSG can produce a compound having differentbiological/chemical properties and/or activity. Thus, depending on thedesired use of a drug comprising a composite including the oxidizedglutathione-based compound, it is possible to obtain a particular drugfor treatment of a particular disease. In addition, new chemicalmodifications of the oxidized glutathione-based compound, such asaminogroup acylation (for instance, bis-phenylalanyl-glutathione, andetc.), can result in a significant decrease in the risk of secondaryreactions due to disulfide bond destruction. Reactions such asS-alkylation, oxidation to the corresponding acids, etc., can causeparticular hardships in the working process and, in the case given, canbe minimized or excluded.

[0099] The synthetic method of the present invention andphysical-chemical modifications thereof directly in the course of thesynthesis (the disulfide bond and general molecular conformationstabilization; the new reactive functional sites appearance) can resultin obtaining the biologically active composite, such as the platinummaterial and GSSG, having new biophysical, chemical and biochemicalparameters as dictated by the various structural-functional properties.

[0100] In one embodiment, the oxidized glutathione-based compound can bea derivative of glutathione. The glutathione can be derivatized afterpreparation of the composite, or it can be derivatized prior topreparation of the composite, i.e., the GSH can be derivatized prior tooxidation. FIG. 15 depict various examples of derivatives ofglutathione-based compounds, i.e., oxidized glutathione-based compound.FIG. 15a shows the structure of S-thioethylamine•glutathione disulfide.FIG. 15b shows a structure of bis-[DL-6,8-thioetic acid]•glutathionedisulfide. FIG. 15c shows a structure of[β-alanyl-L-histidyl]•glutathione disulfide. FIG. 15d shows a structureof [9-β-D-ribofuranosyladenyl]•glutathione disulfide. FIG. 15e shows astructure of bis-[L-2-amino-4-[methylthio]butanoic acid]•glutathionedisulfide.

[0101] In one embodiment, the oxidized glutathione-based compound has anacylated primary glutamic acid amino group. This variant is mostsuitable for acylation by N-protected activated amino acid derivatives,where after the stage of temporary protection of the cysteinemercapto-groups followed by condensation (activated ester method), andremoval of the N- and S-protective groups and oxidation by the hydrogenperoxide with addition of cis-diamminedichloroplatinum results in theGSSG.Pt composite modified by amino-acids at the glutamic acidaminogroups. In this embodiment the oxidized glutathione-based compoundcan be selected from the group consisting of bis-[methionyl]•glutathionedisulfide (FIG. 16a), bis-[aspartyl]•glutathione disulfide (FIG. 16b),bis-[histidyl]•glutathione disulfide (FIG. 16c),bis-[3-iodine-tyrosyl]•glutathione disulfide (FIG. 16d),[γ-aminobutanoyl]•glutathione disulfide (FIG. 16e),bis-[γ-hydroxybutanoyl]•glutathione disulfide (FIG. 16f),bis-[DL-6,8-thioetic acid]•glutathione disulfide (FIG. 15b), andbis-[3,4-dihydroxyphenylalaninyl]•glutathione disulfide (FIG. 16g).

[0102] When the composite includes phenylalanyl groups, the compound isbis-phenylalanyl-GSSG.Pt (see FIG. 7). According to this basic scheme(as shown in FIG. 22), all other GSSG.Pt modifications based on theaminogroups acylation by the derivatives of the protected aminoacids,oxy-acids, carbonic acids and derivatives thereof are synthesized.Slight changes of the methods are possible due to a specific nature ofmodifying molecule. Protecting groups for the initial GSH compoundinclude Bam (N-hydroxymethylbenzamide). The amino acid (AA) can beprotected with groups such as BOC (butyl carbamate) or O—Su(N-oxysuccinimide ester; see FIG. 8 for specific details).

[0103] When the composite is bis-methionyl-GSSG.Pt, i.e., (Met)₂GSSG.Pt,incorporation of a methionine group may involve BOC-protective groupdeblocking after the condensation stage.

[0104] When the composite is bis-aspartyl-GSSG.Pt, an aspartic acidgroup can be introduced according to the general aforementioned scheme.A temporary protective group for the β-carboxyl group is preferablyremoved, if possible, with the BOC-protective group simultaneously ifneeded. A β-tert-butyl ester of N-BOC-aspartic acid, i.e.,BOC-Asp(OBu^(t))-OSu, can be applied at the condensation stage. Theprotective groups can be removed by trifluoroacetic acid (TFA).

[0105] When the composite is bis-histidyl-GSSG.Pt, a histidine imidazolering can be protected via a di-tert-butyl-oxycarbonyl histidinederivative, i.e., BOC-His(BOC)-OSu, which can be used at thecondensation stage. As in previous cases, the protective groups can beremoved by trifluoroacetic acid.

[0106] When the composite is bis-3-iodine-tyrosyl-GSSG.Pt, a possibleprotecting group is an acid-labile protective group such as tert-butylester. The tyrosine derivative used at the concentration stage can beBOC-Tyr(OBu^(t))-OSu.

[0107] When the composite is GABA-GSSG.Pt, (γ-aminobutanoyl-GSSG.Pt), anacid-labile protective tert-butyloxycarbonyl (BOC) group can be used atthe condensation stage.

[0108] When the composite is GOBA-GSSG.Pt, (γ-hydroxybutanoyl-GSSG.Pt),an acid-labile group tert-butyl ester (which can be removed by atrifluoroacetic acid solution) can be used to protect a GOBA hydroxylgroup. A derivative, used at the condensation stage, can beGABA(OBu_(t))-OSu.

[0109] When the composite is bis-lipoyl-GSSG.Pt, it is believed thatside functional groups of lipoic acid do not require special protection.At the condensation stage it is possible to apply an activated(hydroxysuccinimide) ester of lipoic acid. There may be no need for aTFA treatment.

[0110] When the composite is bis-3,4-dihyrooxyphenylalanyl-GSSG.Pt(bis-DOPA-GSSG.Pt), to introduce DOPA molecule, it may be necessary topreviously protect two hydroxyl groups of 3,4-dihydroxyphenylalanine bytert-butyl esters and to protect the aminogroup by a BOC-protectivegroup. For condensation with a composite precursor, an activated estercan be obtained (hydroxysuccinimide or pentafluorophenyl one) that isused in excessive mole amount. Removal of the protecting groups canoccur simultaneously with a trifluoroacetic acid solution.

[0111] When the composite is bis-[carnosyl]-GSSG.Pt(bis-β-alanyl-L-histidyl-GSSG.Pt), carnosine can be protected as adi-BOC derivative of the β-alanine aminogroup and histidine imidazolegroup before condensation. Then condensation and following deblockingcan be performed as previously described. A structure ofbis-[carnosyl]-GSSG can be found in FIG. 19a.

[0112] In another embodiment, the oxidized glutathione-based compoundhas an amide or phosphoramide bond to a unit selected from the groupconsisting of heterocyclic carbonic acids and nucleotides. In thisembodiment, examples of the oxidized glutathione-based compound includebis-nicotinoyl-glutathione disulfide(bis-[pyridine-3-carbonyl]•glutathione disulfide) (FIG. 17a),uridine-5′-monophosphatoyl•glutathione disulfide (FIG. 17b),inosine-5′-monophosphatoyl•glutathione disulfide (FIG. 17c),folliculylsuccinyl•glutathione disulfide (FIG. 17d) andglycerol-1,3-diphosphatyl•glutathione disulfide (FIG. 17c).

[0113] When the composite is bis-nicotinoyl-GSSG.Pt(bis-pyridine-3-carbonyl-GSSG.Pt), a nicotinic acid containing no sidefunctional groups can be introduced into condensation with a compositeprecursor without obtaining protected derivatives as correspondingactivated esters such as hydroxysuccinimide or pentafluorophenyl. TFAtreatment for the removal of protecting groups may not be required.

[0114] When the composite is uridine-5′-monophosphatoyl-GSSG.Pt(UMP-5′-GSSG.Pt), uridine-5′-monophosphate in presence ofN,N-dicyclohexyl-carbodiimide can form phosphoamide links in reactionswith amides (Chambers R. W., J.Am.Chem.Soc., 80, 3752, 1958). Thecomposite precursor can have protected carboxyl groups, such astetratrimethylsilyl derivatives to be used as an aminocomponent.Deblocking can proceed in mild water-alcohol systems.

[0115] When the composite is ionosine-5′-monophosphatoyl-GSSG.Pt,IMP-5′-GSSG.Pt, the synthetic scheme would most likely be similar tothat of the previous derivative (UMP-5 ′-GSSG).

[0116] When the composite is folliculylsuccinyl-GSSG.Pt, a link betweenGSSG.Pt and estrone can be made by amide and ester bonds through asuccinyl residue. Estrone can be transformed into an activatedderivative by reaction with succinanhydride with following condensationby N,N-dicyclohexylcarbodiimide with a protected or blocked compositeprecursor and with tetra-trimethylsilyl derivatives as well. Deblockingcan be performed in a water-alcohol system.

[0117] When the composite is glycerol-1,3-diphosphatyl-GSSG.Pt, themodification can proceed by a carbodiimide method using a compositeprecursor protected, as a tetra-trimethylsilyl derivative as an aminocomponent, the synthesis is similar to that in the synthesis ofphosphoamide derivatives (See Examples 13 and 14).

[0118] In another embodiment, the oxidized glutathione-based compoundcan be selected from the group consisting of tetra-dopamine•glutathionedisulfide (FIG. 18a) and theophylline•glutathione disulfide (FIG. 18b).The formation of amide links can occur between composite carboxyl groupsand amides. The reactivity of all four carboxyl groups is very similarand, therefore, a mixture of products can result.

[0119] When the composite is tetra-dopamine-GSSG.Pt, a 3,4-di-tert-butylester of dopamine can be used as an aminocomponent anddi-tert-butyloxycarbonyl derivatives the composite can be used as acarboxyl component. Condensation can proceed by aN,N-dicyclohexyl-carbodiimide, and removal of the protective groups canbe performed with a trifluoroacetic acid solution.

[0120] When the composite is GSSG.Pt-theophylline, theophylline can beused as an aminocomponent, a composite precursor can be used for acarboxyl component as a di-tert-butyloxicarbonyl derivative.Condensation can be performed with an “F” complex. The removal ofprotective group can be performed with a trifluoroacetic acid solution.

[0121] In another embodiment, the oxidized glutathione-based compoundcan include mixed disulfides. Possible combined structures can involvemixed disulfide formation (symmetric and non-symmetric). (See FIGS.20a-20 c)

[0122] One structure (FIG. 20b) can be formed via mutual oxidation ofmercaptogroups starting materials. There may be no need for additionalprotective groups and condensation methods.

[0123] Another compound (FIG. 20a) can be obtained by formation of anamide bond between the cyteamine aminogroup and one of the compositeprecursor carboxyl groups. It may be necessary to introduce N-protectivegroups and to activate the composite precursor carboxyl groups. Due tothe presence of four carboxyl groups it may be necessary to manipulatethe stoichiometry and/or perform chromatographic separation of theresulting products.

[0124] The synthesis conditions are different from the structure 20b bypresence of the additional aminocomponent equivalent. At thechromatographic purification the structure 20b is used a witness.

[0125] It is obtained from the structure 20c through formation of anadditional disulfide bond at the mercapto-group reaction. During thechromatographic separation of products, it may be necessary to have thestructure 20c as a witness.

[0126] In another embodiment, the oxidized glutathione-based compoundcan be a salt selected from the group consisting of alkali metal salts,alkaline earth metal salts, and transition metal salts. Examples of suchsalts include divanadate salts (FIG. 21a), dihydrofluoride salts (FIG.21b), dilithium salts (FIG. 21c), didopammonium salts, and dizinc salts(FIG. 21d).

[0127] The salts can be obtained through addition of the correspondingamount of the salt-forming components, a base or an acid. Examples ofsalts with aminogroups includes a divanadate of GSSG.Pt((HVO₃)₂-GSSG.Pt) or a dihydrofluoride of GSSG.Pt ((HF)₂.GSSG.Pt).Examples of salts with carboxyl groups include a dilithium salt GSSG.Pt(see Example 2), a GSSG.Pt didopammonium salt or a GSSG.Pt zinc salt.

[0128] In another aspect of the invention, a drug comprising thecomposite is obtained according to the previously described method, suchas the hexapeptide bis-(γ-L-glutamyl)-L-cysteinyl-bis-glycine disodiumsalt and cis-diamminedichloroplatin (cis-platin). Thus, the presentinvention provides a new class of medicinal substances “thiopoietins”that can be introduced into biological media, resulting in a new levelof metabolism and cellular genetic activity.

[0129] An advantageous feature of the composite of the present inventionis the biological activity, in particular:

[0130] Stimulation/modulation of the endogenous production for asignificantly large range of the cytokine, growth and hemopoieticfactors in conditions of radiation and chemical immunosuppression (IL-1αand β, IL-2, IL-3, IL-4, IL-6, IL-8, IL-10 and IL-12, TNF-α, IFN-α andIFN-γ, erythropoietin, colony-stimulating factors) (See Examples 5-7);

[0131] Reproduction of some cytokine effects (IL-2, IL-12, IFN-α andIFN-γ) due to the mechanism induction for the redox-sensitive expressionof the immunologically significant genes and the key protein “critical”cysteine modification for the cellular signal-transducing systems (SeeExample 8);

[0132] Restoration of the depressed bone-marrow haemopoiesis includingerythrocyte, leukocyte and platelet counts as well as levels of CD3⁺,CD4⁺, CD8⁺, CD16/56⁺, CD19/20⁺, CD25⁺, CD34⁺, CD95⁺ (See Examples 9-13)at patients receiving radiation and high-dose combined chemotherapy;

[0133] In conditions of an active antibacterial, antiviral and antitumorchemotherapy, hepatotropic effects as well as diminution of the cardio-,hephro- and neurotoxicity signs;

[0134] The differentiated impact with regard to normal cells (includingthe ones being under functional stress) and transformed ones, namely,the metabolism, proliferation and differentiation stimulation at thenormal cells/tissues and, simultaneously, the apoptosis mechanisminduction only in the tumor- and/or virus-transformed cells. The dataappears to support the induction of p53-dependent and p53-independent,Bcl-2-intensive apoptosis pathway/types occurring (See Examples 14-16).

[0135] The method for the production of the composite makes possiblesynthesis of different composites as a basis for design of drugspossessing a range of new features, namely:

[0136] Increased biochemical drug stability, i.e., “non-assailability”by the GSSG metabolism enzymes (i.e., far less accessible for theseenzyme action), first of all, by NADP.H⁺-dependent glutathione reductasethat basically increases the drug half-life time in the biological mediaexactly in the disulfide form;

[0137] New biophysical component with high level of the donor-acceptorpotential;

[0138] Presence of new reactive sites within the said molecule and,therefore, entirely new capacity for chemical modification.

[0139] The composite properties allow it to function as a uniquecellular “gyroscope” that in conditions of the extreme externalenvironmental factors (physical, chemical and biological ones) providesrestoration of a balance:

[0140] Within the cytokine profile, i.e., the cytokines regulatingproliferation mainly; and the cytokines regulating mainlydifferentiation of the immunocompetent cells;

[0141] Ratio of the cellular redox potential including donor/acceptorbalance of the electron dynamics due to restoration of thethiol-disulfide metabolism; NAD/NAD.H and NADP/NADP.H ratios;

[0142] cAMP/cGMP ratio, changes of the extra- and intracellular ionizedcalcium;

[0143] Relationship of the transcriptional differentiation factors(NFκB) and proliferation factors (AP-1); relationship of the functionalactivity manifestations of p53, p21 and ras-proteins, therefore, balanceof cellular proliferation, differentiation and apoptosis taking intoaccount basically different exhibitions of these effects in the normaland transformed cells.

[0144] In the applied invention, including examples of the preferredembodiments, the following terminology accepted is used.

[0145] “Subject in need thereof” as used in this application hereincomprises a mammal, e.g., man, domestic animals and livestock includingcats, dogs, cattle and horses, having one or more manifestations of adisease in which stimulation of endogenous cytokine or hemopoieticfactor (or both) production as well as an apoptosis mechanism regulationwould be considered beneficial by those skilled in the art with anup-to-date biomedical knowledge.

[0146] “Medicinal drug” as used in this application includes any drugform containing the composite of the present invention, e.g., GSSG.Ptand derivatives, which has a therapeutic effect on neoplastic,infectious, hematologic, immunologic, neurodegenerative or otherdiseases.

[0147] “Therapeutic effect” indicates any effect in man and othermammals which is beneficial, including curative, preventative, allowingmaintenance at a beneficial level, or being in any way advantageous inregard to a body of man and other mammals.

[0148] “Pharmaceutically acceptable salt” as used in this applicationcomprises any composite derivative in the form of a salt that isacceptable for use in the body without unwanted detrimental effect onthe body, and including, for example, sodium, lithium, zinc or vanadiumcations, or sodium, lithium, zinc or vanadium salt respectively.

[0149] “Pharmaceutically acceptable composition” as used in thisapplication involves the composite, or derivative thereof, as apharmaceutically acceptable substance and may include, in addition, agroup of active metabolites or other chemical compounds. For example,the composite or derivative can include or can be covalently bound tophenylalanine or to cystamine.

[0150] “Metabolism” as used in this application involves the totality ofall biochemical reactions taking place within the living organismresponsible for vital function maintenance in the said organism (RobertC. Bohinski “Modern Concepts in Biochemistry”, 4th edition, 1987).

[0151] “Proliferation” as used in this application involves reproductionor multiplication of similar forms (cells) due to constituting(cellular) elements (Stedman's Medical Dictionary, 26th edition, Moscow,1995, p. 519; Miller-Keane “Encyclopedia & Dictionary of Medicine,Nursing, & Allied Health:, 5th edition, W. B. Saunders Company, 1992, p.1221).

[0152] “Differentiation” as used in this application involves cellchanges including acquisition or possession of features distinguishingfrom an original with the cell conversion from relatively simplefunctions to more complex, specialized functions as is in morphologicaland/or functional heterogeneity incident to the given cellular typethrough the tissue-specific gene expression (A. Horst “Molecular Basisfor Disease Pathogenesis”, Moscow, 1982, p. 125; Brian W J Mahy “ADictionary of Virology”, 2-e Academic Press, 1997, c. 87; Stedman'sMedical Dictionary, 26th edition, Moscow, 1995, p. 179; and Miller-Keane“Encyclopedia & Dictionary of Medicine, Nursing, & Allied Health”, 5thedition, W. B Saunders Company, 1992, p. 424).

[0153] “Apoptosis” as used in this application involves morphologicallydistinguishable forms of genetically programmed, physiological celldeath initiated by extra- or intracellular signals when there areactivated enzymes (e.g., the caspases group) causing destruction (e.g.,fragmentation) of nuclear DNA through intranucleosomic cuts andmorphologically manifested by (1) cell shrinkage; (2) condensation,margination, and fragmentation of chromatin; and (3) retention ofcytoplasmic organelle structure, but loss of positionalinterrelationships; further apoptotic cells or apoptotic bodies formedout of them are engulfed (incur phagocytosis) (Harrison's Principles ofInternal Medicine, 14th edition, p. 511, 1998; Apoptosis: a role inneoplasia, C. D. Gregory, 1996; and The Molecular Biology of Apoptosis,D. L. Vaux, A. Strasser; Proc. Natl. Acad. Sci. USA 93 (1996)).

[0154] “Cytokines” as used in this application comprises peptide-originregulatory compounds produced by the different cell types playing a keyrole in the immune response development, hemopoiesis and differentdisease pathogenesis, performing their effect through gene activation,participating in regulation for all immune system elements(proliferation and differentiation of immune competent cell precursors;antigen representation, antigen-sensitized lymphocyte proliferation,B-lymphocyte differentiation into antibody-producing cells, T-lymphocytedifferentiation into functionally different T-lymphocytes; functions ofmacrophages, neutrophils, eosinophils, mast cells and basophils), aswell as controlling growth, differentiation, apoptotic processes andfunctional activity for different tissue cells (including fibroblasts,chondrocytes, keratinocytes, endotheliocytes, nerve tissue cells andcardiomyocytes) (Harrison's Principles of Internal Medicine, 14thedition, p. 511 1998; Apoptosis: a role in neoplasia, C. D. Gregory,1996).

[0155] As used herein, the terms “neoplastic and infectious disease”,“hemopoiesis and immunity depression of various origin”, and “otherdiseases” mean any neoplastic or infectious disease, any conditionscaused or accompanied by the erythroid or myeloid suppression, or areduction in quantitative or functional immunity parameters, as well asany other disease or pathological condition, in whichstimulation/modulation of the aforesaid cytokine and/or hemopoieticfactor endogenous production as well as apoptosis mechanism inductionwould be considered advantageous by those skilled in the art. Thus,modulating the cytokine and hemopoietic factor endogenous production fora person in need thereof by using the composite with the stabilizeddisulfide bond (e.g., GSSG•cis-platin), which being introducedparenterally also has a new feature of influencing the cytokine profileallowing regulation of the normal cell metabolism, proliferation anddifferentiation processes.

[0156] One embodiment of the present invention provides a method forstimulating endogenous production of cytokines and hemopoietic factors.The method includes the step of introducing to a mammalian body in needof such stimulation an effective amount of a composite comprising theoxidized glutathione-based compound and a metal material for a period oftime to stimulate the endogenous production to obtain a therapeuticeffect. As described previously, the ratio of the oxidizedglutathione-based compound to the metal material is between about 3000:1to about 1:1 where the metal material comprises a metal selected fromthe group consisting of platinum and palladium.

[0157] Therapeutic effect includes alleviation, prevention or curing ofan unwanted body condition and can comprise a process selected from thegroup consisting of regulating proliferation in normal cells, regulatingdifferentiation in normal cells, and inducing apoptosis of transformedcells where the transformed cells can include diseased cells. Thetherapeutic effect includes preventative, alleviation and curing effectsin various diseases.

[0158] “Disease” refers to any unwanted condition of the body including,but not limited to, selected oncological diseases, infectious diseases,immunological diseases, ischemic diseases, neurodegenerative diseases,metabolic diseases, endocrinal diseases, and any other unwanted medicalcondition.

[0159] The composite can be administered by various methods: orally oras a solution form selected from the group consisting of inhalationsolutions, local instillations, eye drops, intranasal introductions, anointment for epicutaneous applications, intravenous solutions, injectionsolutions, and suppositories. Preferably, the glutathione is introducedparenterally or topically. The method is carried out by introducing thecomposite, derivatives or salts thereof to enhance the regulatoryinfluence on stimulating of the cytokine and hemopoietic factorendogenous production; or to induce apoptosis mechanisms in transformedtissues, which provides regulation of metabolism, proliferation anddifferentiation in tissues and achieving a corresponding therapeuticeffect.

[0160] In one embodiment, the composite is administered in a dosage ofbetween about 0.1 mg/kg to about 1.0 mg/kg by body weight. In anotherembodiment, the composite is administered in a dosage of between about 1mg/m² to about 100 mg/m² by body surface. In another embodiment, thedrugs can be applied one or more times a day, by one or more day pulsesor continuous administration until a desired therapeutic effect isachieved.

[0161] In a preferred embodiment, the GSSG.Pt material pharmaceuticallyacceptable derivatives are introduced to the body at a dose from 0.01 to1.0 mg of GSSG.Pt material per kg of body weight for the GSSG.Ptmaterial or salt thereof; or at a dose from 1 to 100 mg per 1 m² of bodysurface and in case when applied epicutaneously/through instillations ata dose from 1 to 100 mg per 1 m² of body surface as well, at least, onceduring each 24 hour period. Also the drug can be continuously injectedor otherwise introduced to the body to have a 24 hour total dosage from0.01 to 1.0 mg per kg of body weight for GSSG.Pt base and salts thereof,and from 1 to 100 mg per 1 m² of body surface during each 24 hourperiod. Preferably, administration and introduction to the body shouldbe carried out until a desired stimulating effect on the cytokine andhemopoietic factor production or the apoptosis induction and, thus, thecellular metabolism, proliferation and differentiation regulation and

[0162] Where the composite is administered as a solution, preferably thesolution has a concentration of between about 1% to about 10% of thecomposite. Preferably, the pharmaceutically acceptable derivatives ofthe GSSG.Pt material for parenteral use is in a pharmaceuticallyacceptable solvent as, for example, an aqueous solution including water,glucose solution, isotonic solutions of sodium chloride, buffered saltsolutions. Other physiological solvents or carriers can be used. Wherethe composite is administered as an injectable form, preferably theinjectable form comprises the composite in a solution in a concentrationof between about 0.01% to about 3.0%.

[0163] For topical application including application for different bodycavities, organic solvents or carriers may be used in the form ofointments, pastes, creams or suppositories.

[0164] The examples of the invention embodiments given below demonstratefeasibility of the invention practical use and confirm itseffectiveness, and also expediency of using these medicinal drugs as aninjectable solution containing 0.01% to 3% of GSSG.Pt base or the saltsthereof with the dosage range from 0.1 to 1.0 mg/kg by body weight orfrom 1 to 100 mg/m² of body surface. In cases when the GSSG.Pt drugs areadministered like inhalation solutions, local instillations, eye drops,intranasal introduction, or an ointment for epicutaneous applications,or suppositories, the recommended concentration range is from 1% to 10%of GSSG.Pt base or the salts thereof.

[0165] The active principle, the composite of the hexapeptide with thestabilized disulfide bond—GSSG.Pt—capable of stimulating/beneficialmodulating the endogenous cytokine and hemopoietic factor production aswell as inducing of transformed cells apoptosis, may be obtained byoriginal, developed by the authors the peptide synthesis techniqueprovided herein. Thereby, the obtained hexapeptide composite (GSSG.Pt)with purpose for subsequent usage in animals and humans is applied as apharmaceutically acceptable GSSG.Pt derivative in an injectable drugform prepared by dissolving of the bulk substance in sterile water forinjections or in any pharmaceutically acceptable solvent with theresultant concentration 0.01-3.0%. For an in vitro use in experimentalsettings, GSSG.Pt or the derivatives thereof may be dissolved insolvents acceptable for performance of corresponding experiments such asculture media, isotonic saline solutions, glucose solutions and thelike.

[0166] Injectable medicinal forms of the GSSG.Pt, salts and compositionsthereof have been tested in animal studies and as well in wide clinicalstudies and pilot trials on sick persons. The drug form for human andanimal use should be prepared under sterile and pyrogen-free conditionswhile exerting every effort to prevent chemical or bacterialcontamination of the medicinal form.

[0167] Using the maximum achievable concentration of the GSSG.Ptmaterial sodium salt solution (10.0%, 100 mg/ml) in water for injections(or in normal saline), and using the maximum tolerable volumesadministered to mice intra-peritoneally (IP, 2.0 ml), intravenously (IV,0.5 ml), and intramuscularly (IM, 0.05 ml), there have been reached theGSSG.Pt dosage levels of 5000 mg/kg (IP), 1350 mg/kg (IV), and 135 mg/kg(IM), i.e. 1000, 270, and 27 times, respectively, have been obtained inmice, exceeding the maximum recommended human dose. In none of the caseseither animal deaths or any toxic signs were observed, proving, in fact,the GSSG.Pt material in injectable drug form are essentially non-toxic.

[0168] Through chemical modifications of the basic GSSG.Pt molecule, itis possible to create new drugs where, in molecules along with the basicstructure that have already demonstrated high medical-biologicalactivity based on the oxidized glutathione composite withcis-diamminedichloroplatinum, there are fragments of othercovalently-bound biochemically active molecules. Usage of the covalentlybound combinations allows enhancing of a range of important drugcharacteristics such as stability and standardized properties ofcomposition. Foundation for the chemical modifications is the GSSG.Pthexapeptide core with two primary glutamic acid aminogroups, cysteinedisulfide bond and carboxyl groups of glycine and a- carboxyl groups ofglutamic acid (FIG. 6).

[0169] Purposely elected covalently-bound fragments of biochemicallysignificant molecules can considerably improve medical-biologicalfeatures of the basic GSSG.Pt material composite making them moreselective for each particular therapeutic purpose, resulting in a sharpincrease in a desirable treatment course. It might be conditioned withan additive effect of fragments in the biochemical activity mechanisms,sharp improvement of the basic molecule transport to a target-cell or atarget-molecule, larger affinity for a receptor, necessaryredistribution of oxidative-reductive (redox) potential and a range ofother factors as well as combination thereof.

[0170] In one embodiment, the drug including the composite of thepresent invention can be useful for the treatment of a variety ofoncological diseases. Included in these oncological diseases are solidtumors including those selected from the group consisting of lungcancer, melanoma, cerebral tumors, colorectal cancer, breast cancer,prostate cancer and ovarian cancer. The drug of the present inventioncan be effective for hematologic tumors (non-solid tumors) includingacute lymphoblastic leukosis and acute myeloblastic leukosis.

[0171] For lung cancer treatment, application of GSSG.Pt is preferredfor the first or fourth stages of the disease as a monoregime. Themonthly course includes dosages from 10 to 100 mg/m² of the body surfaceof intravenous (IV) and intramuscular (IM) drug introduction. In thesecond or third stage, GSSG.Pt can be applied in combination with commonchemotherapy by means of IV, subcutaneous and local (intrapleural)introduction of the drugs. The GSSG.Pt dosage is preferably from 30 to50 mg/m² of the body surface.

[0172] For melanoma treatment, bis-[-iodine-tyrosyl]-GSSG.Pt can beparticularly effective. The monthly course can comprise IV and IM drugintroductions every day, with a dosage of about 30 to 100 mg/m² of thebody surface. Maintenance therapy can occur over a period of up to sixmonths-one year. It comprises subcutaneous (locally, close to theimpaired zones) and IV drug introductions once a week, with a dosage ofabout 10-30 mg/m² of the body surface.

[0173] For treatment of cerebral tumors, bis-[dopamine]-GSSG.Pt ispreferred and may be applied by means of IV injections andcatheterization of the carotid artery. The dosage is preferably about 10to 100 mg/m² of the body surface. The IV introduction can last threeweeks, once every other day. For introduction through the carotidartery, application preferably occurs twice every day during six toseven days. The repeat course can be administered two weeks later, overa period of 21 days as well and comprise IV injections mainly.

[0174] For treatment of colorectal cancer, i.e., adenocarcinoma ofstomach and pancreas, bis-[cystamine]-GSSG.Pt is preferred and can beapplied by means of IV and subcutaneous injections with a dosage ofabout 10 to 30 mg/m² of the body surface. Two to three weekadministration can be used as a pre-surgery preparation and post-surgerypatient management. In case of inoperable stomach cancer, applying thedrug through a fibergastroscope is recommended, introducing the drugaround the cancer infiltrate once every week with the single drug doseof 30-100 mg in this case. Preferably, the solution volume does notexceed 3 mL.

[0175] For breast cancer treatment, cystamine-GSSG.Pt is preferred bymeans of IV and subcutaneous injections with a dosage of about 10 to 100mg/m² of the body surface as a pre-surgery preparation and post-surgerypatient management. Maintenance therapy can take up to one year and cancomprise application of cystamine-GSSG.Pt along with ordinarychemotherapy by courses lasting up to two weeks. There can be one-monthintervals between courses.

[0176] For prostate cancer treatment, application of the zinc salt ofGSSG.Pt is preferred through IV injections and through the drugintroduction into lymphatic spaces with a dosage of about 10 to 100mg/m² of the body surface. The approximate treatment course duration canoccur over a period of three to seven weeks.

[0177] For ovarian cancer treatment, application of theophylline-GSSG.Ptis preferred as an anticancer medicine and as a remedy that restores thecancer susceptibility to cis-platin. There can be IV and endolumbalroutes for the drug introduction with a dosage of about 10 to 100 mg/m²of the body surface over a period of three to seven weeks. It ispossible to combine this treatment with antiestrogen therapy.

[0178] For treatment of acute lymphoblastic leukosis, lithium salt ofGSSG.Pt is preferably applied intravenously as a month course with adosage of about 10 to 50 mg/m² of the body surface along with ordinarychemotherapy.

[0179] For treatment of acute myeloblastic leukosis, the lithium salt ofGSSG.Pt is preferably applied in combination with cystamine-GSSG.Ptthrough IV introduction with a dosage of about 10 to 100 mg/m² of thebody surface. The duration of treatment can occur over a period of aboutthree weeks. The maintenance treatment can be repeated after three-monthperiods during one year with a treatment course duration of about 14-17days.

[0180] Infectious diseases that can be effectively treated with the drugof the present invention include tuberculosis, viral hepatitis B and Cand mixed infections (HBV and HCV), herpes, meningitis (sepsis),peritonitis, acute pancreatis and supporative post-surgery sequalae.

[0181] The treatment of tuberculosis can involve a disseminated processwith destructive pathologic changes (cavities +) and bacterial discharge(BK +). Preferably, bis-[histidyl]-GSSG.Pt is applied by means of IV andIM injections during one month with a dosage of about 3-10 mg/kg of thebody weight. For the following two months, the dosage can be 1-5 mg/kgof the body weight. The drugs can be introduced intravenously twice aweek or intramuscularly every other day.

[0182] For the treatment of viral hepatitis B and C and mixed-infections(HBV and HCV), GSSG.Pt and inosine-5-monophosphatyl-GSSG.Pt(IMP-5-GSSG.Pt) is preferably applied by means of IV and IM injectionswith daily dosage of 30 and 40-50 mg, respectively. In the case ofhepatitis B (HBV), the treatment course duration can be up to one month.In the case of hepatitis C (HCV) and mixed infections, the treatmentcourse duration is preferably not less than three months with 10-12 dayintervals after each month of the treatment.

[0183] For the treatment of herpes, the course therapy preferablyinvolves GSSG.Pt and IMP-5-GSSG.Pt in a manner similar to that forhepatitis C (HCV) patients.

[0184] For the treatment of meningitis, sepsis, tetra-dopamine-GSSG.Ptis preferably applied by means of IV and IM injections with a dailydosage of about 3-10 mg/kg of the body weight. In meningitis patients,applying intralumbal injections is recommended in dosage of 30-70 mgonce every three to four days until the patient's clinical state, bloodand liquor indices restore to normal.

[0185] For the treatment of peritonitis, GSSG.Pt andtetra-dopamine-GSSG.Pt is preferably applied by means of IV and IMinjections with a daily dosage of about 30-70 mg over a period of 10-14days until there is full restoration to norm of the patient's clinicalstate and objective indices (blood and urine analyses; “liver”biochemistry).

[0186] For the treatment of acute pancreatitis, GSSG.Pt andIMP-5-GSSG.Pt is preferably applied by means of IV and subcutaneous(along a left costal arch) injections with a daily dosage of about 3-10mg/kg of the body weight, preferably every day during the first week andthen three times a week during the following 14-17 days. This treatmentcan result in the restoration of the patient's clinical state to normal.

[0187] For the treatment of supporative post-surgery sequalae,preferably GSSG.Pt and IMP-5-GSSG.Pt is applied in a manner similar tothat for peritonitis.

[0188] The drug of the present invention can be effective for variousimmunological diseases. Such diseases include immunosuppressive diseasessuch as AIDS and immunosuppressions of infectious diseases of radiationor toxic origin. Autoimmune diseases include glomerulonephritis,rheumatoid arthritis, collagenosis, systemic lupus erythematosus, anddiabetes types I and II. Other immunological diseases include atopicforms of allergic conditions which include allergic rhinitis, atopicdermatipis, bronchial asthma and urticaria.

[0189] For the treatment of AIDS, GSSG.Pt anduridine-[5-monophosphatyl]-GSSG.Pt (UMP-5-GSSG.Pt) are preferablyapplied by means of IV and IM injections alternated every other day witha daily dosage of about 1-3 mg/kg of the body weight during 30 days. Thetreatment course total duration can be up to six months with two tothree week intervals after each month of the therapy application. Thistreatment can be combined with common antiviral therapy. In the case ofsuch a combination, the antiviral remedies can be used as “impulse”,i.e., short-term courses (7-10 days for each course). In case of theAIDS-associated encephalopathy, using GSSG.Pt and UMP-5-GSSG.Pt isrecommended in single doses of about 30 to 70 mg, respectively, onceevery week, during one month.

[0190] For treatment of immunosuppressions of infectious diseases ofradiation or toxic (chemical) origin, GSSG.Pt and UMP-5-GSSG.Pt ispreferably applied by means of IV and IM injections with a dosage ofabout 1-3 mg/kg of the body weight, every day, during 10-12 days untilhemopoiesis is corrected to normal and the immune system is restored.

[0191] For the treatment of glomerulonephritis, GSSG.Pt and lithium saltof GSSG.Pt is preferably applied by means of IV and IM injections with adaily dosage of about 10-30 mg, 1-2 times every day during first twoweeks. Subsequent treatments preferably involve applying IM injectionsonly with a dosage of about 30 mg, once every day during one month. Thefull course duration can be up to three months with two week intervalsafter each month.

[0192] For the treatment of rheumatoid arthritis, GSSG.Pt and lithiumsalt of GSSG.Pt is preferably applied by means of IV and subcutaneousinjections close to impaired joints with a dosage of about 10 mg, twotimes every day during three weeks. Preferably, subsequent treatmentsinvolve applying subcutaneous injections only, one injection two tothree times a week during three months.

[0193] For the treatment of collagenosis, the treatment coursepreferably involves GSSG.Pt and lithium salt of GSSG.Pt in a mannersimilar to that of rheumatoid arthritis. The treatment can also involvethe addition of no greater than 500 mg of vitamin C per day.

[0194] For the treatment of systemic lupus erythematosus, the treatmentcourse preferably involves GSSG.Pt and lithium salt of GSSG.Pt in amanner similar to that of collagenosis.

[0195] For the treatment of atopic forms of allergic conditions(allergic rhinitis, atopic dermatitis, bronchial asthma, urticaria),GSSG.Pt and dihydrofluoride-GSSG.Pt [(HF)₂.GSSG.Pt] is preferablyapplied by means of IM, subcutaneous injections and nasal drops with adosage of about 0.1-1 mg/kg of the body weight, two times every dayduring three weeks. No less than two or three treatment courses ispreferred, usually in spring and late fall.

[0196] For the treatment of diabetes-type I, vanadium salt of GSSG.Pt(divanadate-GSSG.Pt) is preferably applied by means of IV and IMinjections with a dosage of about 1-2 mg/kg of the body weight, alongwith bis-[nicotinoyl]-GSSG.Pt with a dosage of about 1-3 mg/kg of thebody weight. Preferably, three to four week treatment courses areperformed every three months.

[0197] For the treatment of diabetes-type II, bis-[lipoyl]-GSSG.Pt ispreferably applied by means of IV and IM injections with a dosage ofabout 3-7 mg/kg of the body weight, along with bis-[nicotinoyl]-GSSG.Pt,dosage -1-3mg/kg of the body weight (mainly within the droppercomposition based on a 0.5% solution of glutamic acid). The treatmentcourses can occur over a period of one month, two to three times a year.

[0198] The drug of the present invention can be effective for thetreatment of ischemic diseases such as ischemic cerebral conditionsincluding post-insult conditions (e.g., infantile cerebral paralysis)and ischemic heart diseases such as those manifested mainly as asyndrome of conduction impairment and arrythmias (tachyarrythmia,bradyarrythmia, and impairment of ventricular conduction due to blockageof the His bundle or branches) and diseases manifested mainly as asyndrome of functional myocardial failure (cardiomyopathies of differentorigin, metabolic myocardio dysfunctions, and post-infarctionconditions).

[0199] For the treatment of ischemic cerebral conditions includingpost-insult ones (e.g., infantile cerebral paralysis),bis-[phenylalanyl]-GSSG.Pt is preferably applied by means of IV and IMinjections with a dosage of about 1-7 mg/kg of the body weight, astreatment courses last three to four weeks and intervals after eachcourse up to one month. The total treatment duration can take up to twoyears.

[0200] For the treatment of ischemic heart disease (IHD) manifestedmainly as a syndrome of conduction impairment and arrhythmias(tachyrhythmia, bradyrhythmia, impairment of ventricular conduction dueto block of the His's bundle or branches), bis-[carnosyl]-GSSG.Pt(bis-β-alanyl-L-histidyl-GSSG.Pt) is preferably applied by means of IVand IM injections with a dosage of about 2-5 mg/kg of the body weight astreatment courses lasting three to four weeks.

[0201] For the treatment of IHD manifested mainly as a syndrome offunctional myocardial failure (cardiomyopathies of different origin;metabolic myocardial dysfunctions, post-infarction conditions),glycerol-[1,3-diphosphatyl]-GSSG.Pt is preferably applied by means of IVand IM injections with a dosage of about 3-7 mg/kg of the body weight,mainly within the dropper composition containing a 5% glucose solution.The treatment courses duration can occur over a period of two to threeweeks, two to three times a year.

[0202] The drug of the present invention can be effective for thetreatment of neurodegenerative diseases such as Alzheimer's disease,hereditiary (Huntington's) chorea, amnyotrophic lateral sclerosis,neuro-AIDS and demyelinating diseases such as multiple sclerosis.Neurodegenerative disease can also include neurobehavioral diseases suchas diseases caused by drug (narcotic) abstinence and behavioral diseasescreated with psychotropic and nootropic drugs, such as cerebral hypoxia(post-ischemic conditions), manic-depressive psychosis andschizephrenia.

[0203] For the treatment of neurodegenerative diseases, such asAlzheimer disease, hereditary (Huntington's) chorea, amyotrophic lateralsclerosis, neuro-AIDS, bis-[3,4-dihydroxyphenylalanyl]-GSSG.Pt ispreferably applied by means of IV, subcutaneous (along cervical-thoracicspine) and IM injections with a dosage of about 1-5 mg/kg of the bodyweight, as treatment courses lasting up to one month during one year.Intervals between courses can be two or three weeks.

[0204] For the treatment of demyelinating diseases, such as multiplesclerosis, bis-[3,4-dihydroxyphenylalanyl]-GSSG.Pt is preferably appliedby means of IV and subcutaneous (along the spine and endolumbal route)injections with a dosage of about 1-10 mg/kg of the body weight, astreatment courses lasting up to one month. The total treatment durationcan last up to one to two years. The drug can be introduced throughendolumbal route one to two times a month.

[0205] Psychotropic and nootropic drugs can be used for treatment ofneurodegenerative diseases such as cerebral hypoxia (post-ischemicconditions), γ-hydroxy-[butanoyl]-GSSG.Pt can be applied by means of IVand IM injections with a dosage of about 1-4 mg/kg of the body weight astwo to four week treatment courses. Introduction of the drugendolumbally is recommended once a week, with a dosage of about 1 mg/kgof the body weight.

[0206] For the treatment of manic-depressive psychosis, such asschizophrenia, γ-amino-[butanoyl]-GSSG.Pt is preferably applied by meansof IV and IM injections with a dosage of about 3-5 mg/kg of the bodyweight as four to six week treatment courses two to three times a year.

[0207] The drugs of the present invention can be effective indiminishing the contents of pre-β- and β-lipoproteins in blood for thetreatment of atherosclerosis and other metabolic diseases.Bis-[nicotinoyl]-GSSG.Pt (bis-pyridine-3-carbanoyl-GSSG.Pt) can beapplied by means of IV and IM injections with a dosage of about 1-3mg/kg of the body weight as treatment courses lasting two to three weeksthree times a year.

[0208] The drugs of the present invention can be effective for thetreatement of endocrinal diseases such ashypothalamic-hypophysial-ovarian associated functional link impairmentsthat can produce hormonal abnormalities and cause sterility.Folliculyl-[succinyl]-GSSG.Pt can be applied by means of IM andsubcutaneous injections with a dosage of about 3-10 mg/kg of the bodyweight, as monthly treatment courses three to four times a year.

[0209] Other preferable derivatives include the GSSG.Pt materialderivatives in the form of its sodium, lithium, potassium, calcium,zinc, molybdenum, vanadium and other salts, as well as the GSSG.Ptderivatives obtained through covalent binding to phenylalanine, or tomethionine and some other aminoacids including D and L forms of theaminoacids herein; or to cystamine, lipoic acid, or to inosine.

[0210] In one embodiment, manifestation of the immunological,biochemical and molecular-biological effects of the GSSG.Pttherapeutical impact can be obtained in the case when a combinationcomprising 50% of GSSG.Pt with all aminoacids in L-form and 50% ofGSSG.Pt with two chemically equal aminoacids being represented in D-formand others being represented in L-form is used.

[0211] The present invention presents the advantageous feature that thedrug comprising the composite or derivatives thereof has a regulatingeffect on the endogenous cytokine production processes and, thus, on theproliferation and differentiation processes of the T-and B-lymphocytesubpopulations (CD⁺-cells). Drug induction can result in production of awide cytokine and hemopoietic factor range and CD⁺-lymphocytes.Therefore, in this range, from the point of view of cytokineinteraction, there are both agonist-cytokines and antagonist-cytokinesregarding the effects they stimulate (e.g., “relationship” of IL-1α andβ and IL-4). In connection with that, depending on the initial patient'simmunogenesis system state, hyper- or hypoactivity, the drugs of thepresent invention can restore a disturbed balance in the system.

[0212] The given provision is illustrated in Examples 9-13 which showthat patients with depressed immunity (oncological patients receivingradiation or combined chemotherapy) the cytokine synthesis induction(IL-1α and β, IL-2, IL-3, IL-4, IL-6, IL-10 and IL-12, IFN-α and IFN-γ)can be accompanied by restoration of CD3⁺, CD4⁺, CD8⁺, CD16/56⁺, CD25⁺,CD34⁺counts; and patients with the immunoautoagression signs—at theclones of cytotoxic lymphocytes or fibroblasts in case of viralhepatitis C the Fas-Ag (CD95⁺) is expressed that promotes the apoptosismechanism induction and elimination of the virus-transformed and/or“aggressive” cells.

[0213] Another advantageous feature of the present invention involvesthe finding of the composite impact on the isolated human lymphocytes 10minutes (a peak is observed at the 30^(th) minute (the maximal level ofphosphorilating of the cytosol proteins obtained from the lymphocytes))after parenteral introduction of GSSG.Pt material, a significantincrease of the phosphorylating level on tyrosine for lymphocytecytosole proteins that is an integrative characteristic for the cellularsignal-transducing system activity. These changes in state for keyfactors of cAMP, cGMP, inositol-phosphate-dependent signal systems owingto the GSSG.Pt material influence (See Example 8) calls forth theredox-sensitive gene expression, first of all, for the immunologicallysignificant genes responsible for the cytokine and hemopoietic factorsynthesis. Therefore, the GSSG.Pt material application in the treatmentpurposes not only stimulates the cytokine and hemopoietic factorendogenous production but also provides reproduction of the biochemicaland physiological cytokine effects, in particular, in the case ofsensitivity loss of receptors to cytokines that is observed atoncological and retroviral pathology.

[0214] In the tumor- and/or virus-transformed cells the apoptosismechanisms are induced through the GSSG.Pt materialmulticytokine-activating impact, its influence on p53-dependent andp53-independent apoptosis induction mechanisms as well as throughchanging of the donor/acceptor π-electron balance in malignant (cancer)cells (see Examples 14-16).

[0215] Depending on the initial patient's biological status includinghis immunity condition: immunodeficiency, i.e., hyporeactivity; orimmunoautoaggression, i.e., hyperreactivity; presence of the tumor- orvirus-transformed cells—the composite and/or pharmaceutically acceptablederivatives thereof are able to act as the endogenous cytokineproduction stimulators/modifiers and/or as the apoptosis mechanisminducers, respectively.

[0216] These compounds and the drug forms thereof obtained which includethe GSSG.Pt material composite are applied as medicinal drugs capable inthe therapeutic purposes depending on the initial subject's biologicalstatus of the subject in need thereof to stimulate/modulate the widerange cytokine and hemopoietic factor endogenous production and/or toreproduce the cytokine effects as well as to perform the differentiatedeffect regarding the normal (the metabolism, proliferation anddifferentiation regulation) and the transformed cells (the apoptosismechanism induction). “Transformed cells” refers to tumor- and/orvirus-transformed cells.

[0217] Performed experimental and clinical investigations show thattherapeutic effects of the drugs obtained from the GSSG.Pt material andderivatives thereof are based on the multicytokineactivating action andcapacity to reproduce cytokine and hemopoietie factor effects. At thesame time, the data was obtained indicating the GSSG.Pt material has adirect antitumor effect, especially regarding GSSG.Pt material saltsadministered in pharmaceutically acceptable drug forms. Moreover, theGSSG.Pt material effect has revealed to be different for normal andtumor cells. The research with use of the normal and tumor cellsdemonstrated that the GSSG.Pt material or the GSSG.Pt material inpharmaceutically acceptable compositions initiated tumor cell deaththrough apoptosis mechanism. In case of normal cells, they did notundergo destruction (See Examples 14-16).

[0218] Also high effectiveness of the applied medicinal drugs based onthe GSSG.Pt material in regard to apoptosis mechanism induction in thevirus-transformed cells, for example, in case of viral hepatitis B andC, should be noted as exemplified in Examples 11-13.

[0219] Therapeutic effects of GSSG.Pt material and pharmaceuticallyacceptable derivatives thereof, particularly, salts thereof for thetreating of oncological, infectious (viral) diseases can be explained asa stimulation of the wide-ranged endogenous cytokine production with aunique ability to activate apoptotic death of the transformed cellsexclusively. Moreover, the majority of the GSSG.Pt therapeutical effectsand the pharmaceutically acceptable derivatives thereof in theexperimental and clinical conditions can be applied to be connected withrevealed properties of the GSSG.Pt material and the drug forms thereofto stimulate/beneficially modulate the endogenous cytokine production orto reproduce their effects in regard to stimulation of the normal cellproliferation and differentiation and, at the same time, to activateapoptotic death of the transformed cells exclusively.

[0220] Another advantageous feature of the drugs of the presentinvention is a correcting influence of the GSSG.Pt material and thesalts thereof on the metabolic abnormalities, particularly, on theimpairment of carbohydrate metabolism at diabetes, type II. In this case(See Example 7) restoration of the normal cAMP/cGMP ratio as well as thethiol-disulfide ratio in tissues due to the GSSG.Pt material impact(vanadium salt thereof) provided stable setting to normal values for theglucose content in the patient blood, which is a considerabletherapeutic effect.

[0221] Resuming the results of performed experimental, preclinical andclinical studies of the medicinal remedies group developed on theGSSG.Pt material one should emphasize that the parenteral (intravenous,intramuscular, subcutaneous, instillations into urinary bladder or perrectum, etc.) introduction of the indicated medicinal remedies provides:a) stimulation/beneficial modulation of the endogenous production ofIL-1α and β, IL-2, IL-3, IL-4, IL-6, IL-8, IL-10 and IL-12, TNF-α, IFN-αand IFN-γ, erythropoietin, G-CSF, M-CSF and GM-CSF; and, thereupon, widerange of biochemical and immunological effects; b) reproduction of theeffects of the said cytokines and hemopoietic factors in case ofcytokine receptor desensitization; as well as—c) induction of apoptosismechanism in tumor- or virus-transformed cells only, calling forth inthe organism of the subject in need thereof the correspondingtherapeutic effect.

[0222] The beneficial effects of the drug of the present invention canbe illustrated in FIGS. 25-27. FIG. 25 delineates the humoral andcellular responses to the drugs comprising the composites of the presentinvention. FIG. 26 depicts the components of the immune system activatedby the drugs of the present invention, the components includingplatelets, white blood cells, red blood cells, cytokines,erythropoietines, T-lymphocytes, neutrophils, monocytes, macrophages,natural killer cells, and β-lymphocytes. FIG. 27 depicts the types ofcytokines stimulated by the drugs of the present invention, thecytokines including IL-1, IL-2, IL-4, IL-6, IL-8, IL-10, IL-12, alphaand gamma interferons, TNF-alpha and GM-CSF. Thus, the novel method forobtaining the composite comprising the GSSG.Pt material the newmedicinal substance (thiopoietins) class is applied and proved, in thattherapeutical effects are determined by firstly found and previouslyunknown properties of performing stimulation of cytokine and hemopoieticfactor endogenous production and their effect reproduction, thereby,performing stimulation and/or modulation of the normal cellproliferation and differentiation as well as inducing apoptosismechanisms in virus- and tumor-transformed cells.

[0223] The presence of a chemical interaction between the disulfide bondof the oxidized glutathione-based compound and the platinum material mayplay a biophysical role in the electron balance of biological systems.While not wishing to be bound by any theory, the chemical interactioncan be loosely thought of as a donor/acceptor pair where the loneelectron pairs on the sulfur atoms can potentially interact with anelectron-deficient material, such as the platinum material. Again, whilenot wishing to be limited by any mechanism, if cellular activity andcellular physical state are determined, at least in part, bydonor/acceptor interactions throughout a biological system, then abalance among electron donors and acceptors having equal biopotentialscan be a life parameter. Such balance alteration can be used forregulation of different cell functions and physical properties(Szent-Gyorgyi A., Proc. Natl. Acad. Sci. U.S., 58,2012 (1967);Introduction to a Submolecular Biology, Academic Press, New York(1960)). Sources of mobile electrons can include π-electrons such aslone-pair electrons of nitrogen, oxygen, and sulfur. There are fewacceptor groups (e.g., —C═O-groups) in a normal cell which can bebalanced via donor/acceptor dynamics of donor electrons such asπ-electrons.

[0224] Continuing along with this non-limiting theory, the malignantcell can be characterized as having dramatic disturbances of thedonor/acceptor balance towards an excess of donors electrons. Acceptormolecules are almost absent in cancer cells. A possible solution to thisimbalance in this situation is the presence of molecules that possessdonor/acceptor features within the same molecule, such as a GSSG.Ptmaterial. Introducing GSSG.Pt into biological media can cause arestoration of the electronic balance in the biological media. Thisrestoration can involve a catalysis by the platinum atom in a reactioninvolving the formation of an active oxygen form, e.g., superoxide anionradicals, singlet oxygen.

[0225] Another non-limiting theory involving cellular electronic balanceis an interaction of GSSG.Pt material with cell GSH generatingoxidative-reductive, i.e. donor/acceptor pair. If there is a high GSHlevel in cells that is characteristic for tumor-transformed cells with ahigh proliferative impulse, the pro-oxidative, i.e. oxidative,properties of GSSG.Pt material exhibit in the most evident way. In thecase the oxidative stress forms in the tumor cells only causingalteration of the tumor cells mitochondria functions formingintracellular signal for the apoptosis mechanisms induction.

[0226] For the normal but “tired”, “exhausted” cells, there may be anoxidative-reductive potential optimization, bioenergetic supply for themetabolic transformations, redox-sensitive adequate expression of thegenome functional sites, in particular, the immunologically significantgenes and transcription factors.

[0227] For the transformed cells GSSG.Pt, there may be anincompatibility with vital functions involving chain transfer reactionof π-electrons, disturbance of the mitochondrial oxidative-reductivereactions of electron/protons transfer reactions along the respiratorychain and dislocation of the NADP.H⁺/NADP.H ratio, i.e., forming theintracellular signal for the apoptosis mechanism induction.

[0228] The new GSSG.Pt pharmacokinetics (comparing to GSSG by itself) inblood and tissues (organs) being introduced into biological mediaindicates that the GSSG.Pt molecule is much less available for the GSSGmetabolism enzymes and, first of all, for the NADP.H⁺-dependentreductase, the main enzyme for the GSSG into GSH reduction. Thereupon,the GSSG.Pt half-life time in the disulfide form in biological mediaincreases significantly (See Examples 3 and 4).

[0229] Basically novel pharmacokinetics of the hexapeptide with thestabilized disulfide bond (GSSG.Pt) compared to the structural analog,i.e., the oxidized glutathione (GSSG), provided an optimal manifestationfor the newly determined biological-pharmacological effects.

Examples of the Invention Embodiments

[0230] SYNTHESIS METHOD. 170 g (0.55 mole) of the reduced glutathioneare suspended in 200 ml of water and, along with stirring, 139 ml (0.55mole) of the 4N NaOH solution and 170 ml of 0.05%cis-diamminedichloroplatinum cis-[Pt(NH₃)₂Cl₂] water solution are added.

[0231] The obtained transparent, slightly yellow solution is cooled to18-20° C. and 283 ml of the 3% hydrogen peroxide solution (H₂O₂solution) is added in little portions over a time period of five minuteswith such a speed rate that the reaction mixture temperature will notexceed 22-25° C.

[0232] Thirty minutes after addition of the hydrogen peroxide solution(H₂O₂) the pH is measured, and then the 4N caustic soda solution isadded drop-by-drop to reach pH=5.6-5.8 along with simultaneoustemperature control, which should be within 22-25° C. Then the coolingis taken away and stirring continued at indoor (room) ° C. temperaturefor 30 minutes more.

[0233] Control of the oxidation reaction completeness is performed bythe HPLC assay. A liquid chromatograph for HPLC, type Beckman, Sol.Module 126, Det. 168, with a column Luna Phenomenex ODS 4.6×250 mm, oran identical one, is prepared. To prepare the HPLC mobile phase 20 cm³of acetonitrile and 1 cm³ of freshly distilled trifluoroacetic acid isintroduced into a 1000-cm³ graduated flask, and the volume is increasedup to the mark by the deionized water. The solution is mixed anddegassed by shaking in vacuum.

[0234] Thirty minutes after addition of the entire hydrogen peroxidesolution amount one will check the oxidation reaction completeness bymeans of the highly productive liquid chromatography (HPLC). Theretowith a microsyringe one will take 10 μl of the reaction mixture anddissolve them in 1 ml of the mobile phase (0.1% trifluoroacetic acid:acetonitril, 98:2). 20 μl of the obtained solution is introduced intothe chromatograph Beckman 126 Solvent Module, Diod Array Detector Module168, the column Luna Phenomenex ODS 4.6×250 mm, or the identical.Elution is performed in isocratic regime, 30 min., in the system 0.1%trifluoroacetic acid: acetonitrile, 98:2; the flow-speed rate 1 ml/min,detection at 220 nm, scanning 190-600 nm.

[0235] The retention time in the aforesaid conditions is 5.0+0.5 min forreduced glutathione, 11.0±0.5 min for oxidized glutathione.

[0236] In case if, according to the HPLC data after the standardchromatogram integration, the oxidized glutathione content is less than97%, the stirring is continued in the same regime 30 minutes more andthe HPLC control is repeated.

[0237] In case when the result is equal or exceeds 97%, the reaction isconsidered as completed and one will pass to the reaction solutionFiltration. Thereto, there is used a filter having pore size not largerthan 0.7 μm.

[0238] Weight loss at drying will not exceed 5% at drying to theconstant weight at 100° C. in vacuum (1 mm Hg) above CaCl₂ and P₂O₅.

[0239] The main material content in the ready product by the HPLC datawill not be lower than 98%.

[0240] Thus, the oxidized glutathione as a composite withcis-diamminedichloroplatinum is obtained.

[0241] Appearance: white odorless powder.

[0242] Solubility: soluble in water, 0.9% isotonic solution of sodiumchloride for injections; insoluble in 95% alcohol, chloroform, ether andother organic dissolvents.

[0243] Solution transparency and color: 0.05 g of the drug solution in10 ml of water is transparent and colorless solution.

[0244] PH of 1% solution: 5.0-6.0 potentiometrically, the device ispH/mV/⁰C meter Cole Parmer, model 59003-15 or identical.

[0245] Authenticity:

[0246] a) amino-acid analysis (6 n HCl, 110° C., 20 hrs.),

[0247] glycine—2.0±15%; glutamic acid—2.0±15%; cysteine—2.0±40%;amino-acid analyzer AAA T-339 M Prague or identical.

[0248] b) HPLC—at the outlet time it corresponds to the standard ofbis-(γ-L-glutamyl)-L-cysteinyl-bis-glycine disodium salt.

[0249] Chromatography conditions: device—BECKMAN “Gold NouveauChromatography Data System” Version 6.0, Diod Array Detector Module 126or identical.

[0250] Assay—20 μl of 0.1% drug solution in the mobile phase,chromatography on the column ULTRASPERE ODS 250×4.6 mm with theconverted C₁₈ phase in the isocratic conditions acetonitrile—0.1%trifluorideacetic acid (2:98); flow rate 1 ml/min., detecting at 220 nm,scanning 190-600 nm.

[0251] Purity (main substance content):

[0252] a) at HPLC not less than 98%:

[0253] b) at the amino-acid analysis: not less than 85% (analysisaccording to Section “Authenticity”, Item “a” with an exact weight).

[0254] c) Sodium (Na) content according to the emission spectral methodis 7.0±0.5%.

[0255] d) Platinum (Pt) content according to the mass spectrometricanalysis is 0.032±0.01%

Method for Element Content Determination

[0256] The exact assay weight (about 50 mg) is dissolved in 50 ml ofbidistilled water and the solution is used for the analysis.

[0257] The platinum content is determined quantifiably by the method ofmass spectrometric analysis with inductively bound plasma at the deviceof the PQe model made by VG Elemental, England. The analysis relativeprecision is 5%.

[0258] The other element content is determined quantifiably by themethod of the atomic-emission spectroscopy with inductive bound plasmaon the device of the model TRACE 61E made by Thermo Jarell Ash, USA. Theanalysis relative precision is 5%. Element content, μg/g: Silver (Ag)<1.0 (less than 0.0001%) Aluminum (Al) 2.0 Arsenic (As) <1.0 Barium (Ba)<0.50 Beryllium (Be) <0.05 Calcium (Ca) 7.0 Cadmium (Cd) <0.05 Cobalt(Co) <0.5 Chromium (Cr) 1.7 Copper (Cu) <0.5 Iron (Fe) <1.0 Potassium(K) <2.5 Selenium (Se) <2.0 Magnesium (Mg) <2.5 Manganese (Mn) <0.2Molybdenum (Mn) <0.2 Nickel (Ni) <0.5 Lead (Pb) <0.40 Strontium (Sr) 1.9Titanium (Ti) <0.5 Vanadium (V) <0.5 Zinc (Zn) 0.65 Antimony (Sb) <0.5

EXAMPLE 1 Synthesis ofbis-(L-phenylalanyl-γ-L-glutamyl)-L-cysteinyl-bis-glycine Disodium Salt

[0259] (I.) General drug characteristics.

[0260] 1. Name:bis-(L-phenylalanyl-γ-L-glutamyl)-L-cysteinyl-bis-glycine disodium salt,composite with cis-diamminedichloroplatinum.

[0261] 2. Structural formula—see FIG. 8.

[0262] 3. Gross-formula: C₃₈H₄₈N₈O₁₄Na₂S₂.[Pt(NH₃)₂Cl₂]

[0263] 4. Molecular weight: 950,94 on C₃₈H₄₈N₈O₁₄Na₂S₂ with Pt content0,033%.

[0264] 5. Appearance: white odorless powder.

[0265] 6. Solubility: soluble in water, 0.9% isotonic solution of sodiumchloride for injections; insoluble in 95% alcohol, chloroform, ether andother organic dissolvents.

[0266] 7. Solution transparency and color: 0.05 g of the drug solutionin 10 ml of water is transparent and colorless.

[0267] 8. pH of 0.1% solution: 5.75 (potentiometry).

[0268] 9. Authenticity:

[0269] a) amino-acid analysis (6 n HCl, 110° C., 20 hrs.), (error margin20%, for cysteine—35%), in correspondence: glycine—2.00; glutamicacid—1.92; cysteine—1.81; phenylalanine—2.04.

[0270] b) NMR(¹H)-spectroscopy, according to—“BRUKER” AM 500, 500 MHz,D₂O.

[0271] 10. Purity (main substance content):

[0272] a) At HPLC not less than 97%:

[0273] Device: BECKMAN “Gold Nouveau Chromatography Data System” Version6.0, Diod Array Detector Module 126. Assay—20 μl of 0.1% drug solutionin the mobile phase, chromatography on the column ULTRASPERE ODS 250×4.6mm with a converted C₁₈ phase in isocratic conditions acetonitrile-0.1%trifluorideacetic acid (2:98); flow rate 1 ml/min., detecting at 220 nm,scanning 190-600 nm, PDA functions—Contour Plot, 3D.

[0274] b) At the amino-acid analysis: not less than 85% (analysisaccording to Item 9a with an exact weight);

[0275] c) Thin-layer chromatography is homogenous, analysis is performedat introduction of 5 μl of the 1% drug solution in the band. There areplates Kieselgel 60_(f) (Merck) 10×5 cm, system: n.butanol—aceticacid—water (4:1:1). Development is performed according to the standardmethods—ninhydrine and chlorine\benzidine. R_(f)=0,15;

[0276] d) Sodium (Na) content according to the emission spectral methodis: 4.8%;

[0277] e) Platinum (Pt) content according to the mass spectrometricanalysis is 0.033%.

[0278] 11. Elements detected content, μg/g: Silver (Ag) <1.0 (less than0.0001%) Aluminum (Al) 2.0 Arsenic (As) <1.0 Barium (Ba) <0.50 Beryllium(Be) <0.05 Calcium (Ca) 7.0 Cadmium (Cd) <0.05 Cobalt (Co) <0.5 Chromium(Cr) 1.7 Copper (Cu) <0.5 Iron (Fe) <1.0 Potassium (K) <2.5 Selenium(Se) <2.0 Magnesium (Mg) <2.5 Manganese (Mn) <0.2 Molybdenum(Mo) <0.2Sodium (Na)  48 mg/g Nickel (Ni) <0.5 Lead (Pb) <0.40 Platinum (Pt) 330μg/g Strontium (Sr) 1.9 Titanium (Ti) <0.5 Vanadium (V) <0.5 Zinc (Zn)0.65 Antimony (Sb) <0.5

[0279] Determination method:

[0280] The exact assay weight (about 50 mg) is dissolved in 50 ml ofdouble-distilled water and the solution is used for the analysis.

[0281] The platinum content is determined quantifiably by the method ofmass spectrometric analysis with inductively bound plasma at a PQedevice made by VG Elemental, England.

[0282] The analysis relative precision is 5%.

[0283] The content of other elements is determined quantifiably by themethod of the atomic-emission spectroscopy with inductive bound plasmaon a TRACE 61E device made by Thermo Jarell Ash, USA. The analysisrelative precision is 5%.

[0284] 12. Weight loss at drying: 10% at drying till the constant weightat 100° C. in vacuum (1 mm Hg) above CaCl₂ and P₂O₅.

[0285] (II.) Synthesis method description.

[0286] 1. Process chemical scheme—see FIG. 8.

[0287] 2. Method description

[0288] (III). Product (I) γ-L-glutamyl-L-cysteinyl-glycine in amount of3.07 g (10 mmol) and N-hydroximethylbenzamide (II) in amount of 5.89 g(13 mmol) is dissolved in 30 ml of anhydrous trifluoroacetic acid (TFA)mix at the room temperature during one hr. Then the solvent is distilledoff in vacuum at 40° C., 30 ml of anhydrous ethyl alcohol is added tothe remainder; the solvent is again distilled off in vacuum and theprocedure is repeated two times more. The product is crystallizedthrough grinding in 50 ml of anhydrous diethyl ether, filtered, washedon the filter with 2×20 ml of anhydrous ether and further it is dried invacuum above KOH and P₂O₅. Recrystallization is done from 90% ethanol.Yield—5.50 g (80%). R_(f)=0,43, Kieselgel 60_(f) (Merck) 10×5 cm,system: n.butanol—acetic acid—water (4:1:1).

[0289] (V). Product (III) in amount of 4.40 g (10 mmol) is stirred inthe mixture of 15 ml of distilled water and 25 ml of dioxane; then alongwith mixing, 10 ml (20 mmol) of 2 N NaOH solution is added.

[0290] Then 3.62 g (10 mmol) of phenylalanine N-hydroxysuccinimide ester(IV) is introduced in the reaction mixture and the stirring is continuedduring 12 hrs at room temperature.

[0291] Then the mixture is evaporated in vacuum at 40° C. to dryness.The residue is dissolved in 200 ml of ethyl acetate and washed by 2×20ml of 1 N sulphuric acid, water, sodium bicarbonate (2×50), water andthe organic layer is above the anhydrous chloride calcium. Then ethylacetate sis distilled in vacuum at 40° C. to dryness and the product iscrystallized from ethyl acetate/ether.

[0292] The crystals are separated by filtration and dried in vacuumabove phosphorus pentoxide to constant weight. The product yield(V)—4.88 g (70%). R_(f)=0,80, Kieselgel 60_(f) (Merck) 10×5 cm, system:n.butanol—acetic acid—water (4:1:1).

[0293] (VI). Product (V) in amount of 6.87 g (10 mmol) is dissolved in20 ml of distilled trifluoroacetic acid and the solution is kept at roomtemperature during two hrs. Then the product is precipitated by absoluteether (about 200 ml), filtered and dried in vacuum above KOH to theconstant weight. The product yield—(V) 5.28 g (90%). R_(f)=0.48,Kieselgel 60_(f) (Merck) 10×5 cm, system: n.butanol—acetic acid—water(4:1:1).

[0294] (VII). Product (IV) in amount of 5.87 g (10 mmol) is dissolved in100 ml of a mixture methanol-water (1:1), then 200 ml (10 mmol) ofmercury acetate solution are added and the mixture is stirred at roomtemperature during one hr. Then the hydrogen sulphur flow is spargedthrough the solution during 20 min. controlling precipitation efficiencyat the assay. The mercury sulphide precipitate is filtered; the filtrateis evaporated in vacuum up to volume of 10 ml; then 200 ml of isopropylalcohol is added and the product is crystallized at cooling to 0-4° C.The crystals are filtered, washed with isopropyl alcohol, acetone anddried in vacuum above P₂O₅. The product yield (VII)—3.72 g (82%).R_(f)=0.30, Kieselgel 60_(f) (Merck) 10×5 cm, system: n.butanol—aceticacid—water (4:1:1).

[0295] (VIII). Product (VII) in amount of 4.54 g (10 mmol) is suspendedin 20 ml of water and along with stirring, 5 ml (10 mmol) of 2 N NaOHsolution and then 4.8 ml of 0.05% water solution ofcis-diamminedichloroplatinum (cis-[Pt(NH₃)₂Cl₂]) is added. The solutionis cooled to 18-20° C. and in little portions during about two min., 5.1ml of 3% solution is added at such a rate so that the temperature willnot exceed 22-25° C. Thirty minutes after introduction of the wholehydrogen peroxide amount, the solution pH is measured and its value isbrought to 5.6-5.8 by adding of the necessary amount of 4N NaOHsolution, while the solution temperature is monitored (it is within22-25° C.). Then the stirring is continued without external cooling for30 min. and then the control analysis of the reaction mixture isperformed by HPLC. With that purpose 10 μl is taken out of the reactionsolution and dissolved in 1 ml of the mobile phase. If, according to theHPLC data, the oxidized form content is equal or exceeds 95%, thereaction is deemed as finished. Otherwise, the stirring at the roomtemperature is continued 30 min. more and the HPLC assay is repeated.

[0296] Then the reaction solution is filtered through the filter withpore size not larger than 0.7 μm and the filtrate is lyophilized. Theobtained dry product is dried out in vacuum at 40° C. above anhydrouscalcium chloride to the constant weight. Yield—4.51 g (95%).

[0297] The ready substance is analyzed according to see Example 2.

EXAMPLE 2 Synthesis of bis-(γ-L-glutamyl)-L-cysteinyl-bis-glycineLithium Salt

[0298] (I.) General drug characteristics.

[0299] 1. Name: bis-(γ-L-glutamyl)-L-cystinyl-bis-glycine dilithium saltwith cis-diamminedichloroplatinum.

[0300] 2. Structural formula—see FIG. 9.

[0301] 3. Gross-formula: C₂₀H₃₀N₆O₁₂Li₂S₂ . [Pt(NH₃)₂)₂Cl₂]

[0302] 4. Molecular weight: 624.49 on C₂₀H₃₀N₆O₁₂Li₂S₂ with Pt content0,032%

[0303] 5. Appearance: white odorless powder.

[0304] 6. Solubility: soluble in water, 0.9% isotonic solution of sodiumchloride for injections; insoluble in 95% alcohol, chloroform, ether andother organic dissolvents.

[0305] 7. Solution transparency and color: 0.05 g of the drug solutionin 10 ml of water is transparent and colorless.

[0306] 8. pH of 0.1% solution: 5.0-6.0 (potentiometry).

[0307] 9. Authenticity:

[0308] a) amino-acid analysis (6 n HCl, 110° C., 20 hrs.) incorrespondence: glycine—2.0 (2.0); glutamic acid—1.9 (2.0); cysteine—1.7(2.0).

[0309] b) NMR(¹H)-spectroscopy, in correspondence: CH₂ (δ6 2.05, 2.40,3.00, 3.80); CH (δ 3.72, 4.65).

[0310] c) HPLC corresponds with the standard according to the yieldtime.

[0311] 10. Purity (main substance content):

[0312] a) at HIPLC>95%;

[0313] b) at the amino-acid analysis>85%;

[0314] c) Thin-layer chromatography (TLC) homogenous;

[0315] d) Lithium (Li) content according to the emission spectral methodis 2.2±0.1%

[0316] e) Platinum (Pt) content according to the mass spectrometricanalysis is 0.01-0.02%

[0317] (II.) Staged scheme for the product synthesis (A→B→C)

[0318] A. Oxidation of reduced glutathione(γ-L-glutamyl-L-cysteinyl-glycine)

[0319] A₁—stage for the source reduced glutathione processing byhydrogen peroxide at pH=8—see FIG. 10.

[0320] Source compound (I):γ-L-glutamyl-L-cystcinyl-glycine (GSH)

H-γ-L-Glu-L-Cys-Gly-OH

[0321] Reagents: 1) hydrogen peroxide (35%) Fluka

[0322] 2) ammonia solution (25%)

[0323] Reaction conditions: stirring of the water solution (I) and thereagents at 20° C. (pH=8.0) during 20 min.

[0324] A₂—separation of free hexapeptidebis-(γ-L-glutamyl)-L-cysteinyl-bis-glycine (see FIG. 11)

[0325] Source compound: reaction mixture of the A1 stage.

[0326] Reagents: glacial acetic acid

[0327] Reaction conditions: acidification of the A₁ reaction mixture byglacial acetic acid up to pH=5.0, the solution filtration and lyophilicdrying of the product.

[0328] Control for the A oxidation stage processing: by HPLC at theDelta Pack C18 column (0.1% TFA-MeCN, 0-25%); one will check presence ofthe peak (>97%) for the compound

[0329] (III) (7.4±0.4 min) and the peak absence for the compound (I)(3.0±0.3 min).

[0330] B. Conversion of the oxidized form (III) into the lithium salt(IV)—see FIG. 12.

[0331] Source compound: free hexapeptide (III).

[0332] Reagents: 1 N LiOH solution.

[0333] Reaction conditions:

[0334] a) titration of the compound (III) water solution by two LiOHequivalents;

[0335] b) water evaporation in vacuum at 35-40° C.;

[0336] c) the product precipitation by isopropyl alcohol;

[0337] d) precipitate filtration;

[0338] e) precipitate washing by acetone;

[0339] f) the product drying in vacuum (1 mm Hg) at 35-40° C.

[0340] Control of the B stage processing: the reagents quantities andtechnological conditions at drying is inspected.

[0341] C. Finished product quality control.

[0342] 1. The main material content according to HPLC:>97%.

[0343] The analysis is performed using 20 μl of 0.1% drug solution inthe mobile phase, on the column 250×4.6 mm with a converted C₁₈ phase inisocratic conditions acetonitrile—0.1% trifluorideacetic acid (2:98);flow rate 1 ml/min., detecting at 220 nm. The comparison is made withthe standard peak obtained in the same conditions.

[0344] 2. The main material content according to the spectrophotometricanalysis data on the non-oxidized thiol groups content:>95%.

[0345] Analysis: 0.12 ml of 0.5% drug solution is placed into a 25-mlmeasuring flask, 1 ml of 0.1% Tris-HCl buffer with pH=8, 0.01 M EDTA and1 ml of 2% NaBH₄ solution. The reaction mixture is incubated at 20° C.during 30 min. The reaction is terminated through introduction of 0.6 mlof 1 M HCl during two min. in portions of 0.05 ml along with agitationand following introduction of 2 ml of acetone during three min. withstirring. Then 0.25 ml of the Elman reagent is added and the volume isbrought to the mark by 0.1 M the phosphate buffer solution (pH 8.0). Anoptical density is measured at 412 nm, water is a comparison solution.Simultaneously, the procedure with the drug standard is conducted andthe obtained data is compared.

[0346] 3. Presence of foreign admixtures: according to TLC data the drugis homogenous.

[0347] Analysis is performed at introduction of 5 μl of the 0.1% drugsolution in the band. The plates are Kieselgel 60_(f) (Merck) 10×5 cm,system: n.butanol—pyridine—acetic acid—water (150:100:30:120).Development is performed according to the standard methods—ninhydrineand chlorine\benzidine.

[0348] 4. Lithium (Li) content according to the emission spectral methodis 2.2±0.1%.

EXAMPLE 3 Pharmacokinetics and Metabolism of GSSG.Pt and GSSG in BloodSerum and Tissues After Intravenous Introduction

[0349] The time-concentration GSSG curves and activity changes forenzymes participating at the GSSG metabolism after the GSSG.Pt and GSSGintravenous introduction in different doses were studied. The variationof the oxidized glutathione concentration was evaluated in animal bloodserum, liver, kidneys, spleen and lymphocytes during 60 min. after thesingle GSSG.Pt and GSSG intravenous introduction in doses 2 mg/kg and 20mg/kg of body weight. In addition, the activity variation of the enzymesparticipating in GSSG metabolism was evaluated (glutathione reductase,glutathione-peroxidase,glutathione-S-transferase,γ-glutamyl-transpeptidase).

[0350] The study was performed at male CBA mice (standard bodyweight—180 to 200 g). Five groups of animals (with no less than 15 micein each) were formed. The group description is represented below.

[0351] Control groups:

[0352] #1—intact animals receiving a single injection of the testedarticle vehicle (normal saline—(NS)) instead of the drug;

[0353] Test groups:

[0354] #2—animals receiving the GSSG injection (GSSG dissolved in normalsaline) in a dose of 2 mg/kg;

[0355] #3—animals receiving the GSSG injection (GSSG dissolved in normalsaline) in a dose of 20 mg/kg;

[0356] #4—animals receiving the GSSG.Pt injection (GSSG.Pt dissolved innormal saline) in a dose of 2 mg/kg;

[0357] #5—animals receiving the GSSG.Pt injection (GSSG.Pt dissolved innormal saline) in a dose of 20 mg/kg.

[0358] The blood samples were taken at 1, 2, 5, 10, 20, 40 and 60 min.,the serum was separated and the concentration analysis was performedaccording to a standard method where the main stages are proteinprecipitation, removal from the sample of non-polar and medium-polarcompounds and the following chromatographic analysis withspectrophotometric detection in conditions of isocratic and lineargradient elution.

[0359] The GSSG content variation in the blood serum, different organsand the lymphocytes at the drug intravenous introduction are given inthe Tables 1-4.

[0360] Activity of the enzymes participating in the GSSG metabolism(glutathione reductase: EC.1.6.4.2; glutathione-peroxidase: EC.1.11.1.9;glutathione-S-transferase: EC.2.5.1.18; g-glutamyl-transpeptidase:EC.2.3.2.2) were determined by the standard reagent kits produced byBoehringer Mannheim GmbH. The enzyme activity variation values after theGSSG.Pt and GSSG intravenous introduction in dose of 2 mg/kg are givenat the Tables 5, 6, 7.

[0361] Comparing the drug dynamic distribution of the drugs in the bloodserum, liver, kidneys, spleen and lymphocytes a clear advantage for theGSSG.Pt pharmacokinetics regarding GSSG is evident. The GSSGconcentration to the 10^(th) minute is almost equal to the initial onewhereas, at the same time, the GSSG.Pt concentration exceeds 50 timesthe initial parameters and remains at the given high level till the endof the studied period. Besides, the maximal GSSG.Pt concentration in theblood serum and in the tissues exceeds 3 times the maximal GSSGconcentration. These features determine higher effective duration ofimpact for GSSG.Pt comparing to GSSG.

[0362] As it follows out of the Tables 5, 6 ,7 materials the GSSG drugincreases approximately two times the activity for the enzymesparticipating in the thiol metabolism with large number of provedindices whereas the GSSG.Pt drug does not significantly alter theenzymes activity and the proved indices number is insignificant. Itindicates the greater GSSG.Pt drug stability as a substrate in regard tomain enzymes participating in the glutathione metabolism and, therefore,calls out longer presence of the glutathione oxidized form in the bloodserum and different organs.

[0363] Thus, the obtained data analysis demonstrated the higher GSSG.Ptstability to selective impact of the main glutathione metabolism enzymes(first of all, glutathione-reductase) that determines new dynamics forthe drug pharmacokinetics. It facilitates manifestation of newbiological-pharmacological effects and, thereupon, new GSSG.Pttherapeutic effects.

EXAMPLE 4

[0364] Table A: GSSG pharmacokinetics (initial data)

[0365] A graph on mean values is presented in FIG. 23, showing a GSSGpharmacokinetic curve for intravenous introduction.

[0366] Table B: Pharmacokinetic parameters

[0367] Table C: GSSG.Pt pharmacokinetics

[0368] A graph on mean drug concentration values is presented in FIG.24, showing a GSSG.Pt pharmacokinetic curve for intravenousintroduction.

[0369] Table D: Pharmacokinetic parameters.

EXAMPLE 4 Comparative Analysis of the GSSG.Pt and GSSG Pharmacokineticsat Experimental Intravenous Introduction

[0370] TABLE A GSSG pharmacokinetics Initial data 0 1 2 5 10 20 40 60Assays 1-4 5-8 9-12 13-16 17-20 21-24 25-28 Drug concentrations, 0.246.7 43.15 17.6 7.0 1.5 1.9 5.6 μg/mL 0.6 86.6 31.1 7.0 3.2 0.04 73.623.7 19.1 5.6 3.8 0.1 71 41.3 10.6 7.7 4.8 Ar. mean. 0.3 68.1 47.3 17.310.2 3.8 2.9 1.9 Mean error 0.02 7.6 9.2 3.8 3.0 0.9 1.0 1.8

[0371] A graph on mean values is presented in FIG. 23, showing a GSSGpharmacokinetic curve for intravenous introduction. TABLE BPharmacokinetic parameters Estimation on Direct method Error model ErrorD, mg/kg 10 10 Dose Cmax, 68.1 7.6 72 6.8 Maximal concentration μg/mL m,min 1 1 0.5 0.5 Time for maximum attainment AUC, 60 439 54 446 43 Squareunder the curve before min, μg/mL 60 min. AUC, t min, 455 50 452 47 Fullsquare under the curve μg/mL AUCt/Cmax, 6.5 0.6 6.3 0.7 Effectiveduration min.

[0372] TABLE C GSSG Pt pharmacokinetics Initial data Time, min. 0 1 2 510 20 40 60 Assays 1-4 5-8 9-12 13-16 17-20 21-24 26-28 0.15 265.5 213.698 5 9 1.6 0.8 0.3 186.2 11.95 56.7 19.8 6 1.1 0.3 112.5 87.4 25.8 6.751.1 0.9 29 9.6 4.4 0.4 0.3 Mean concentration, 0.23 187.4 112.8 67.815.1 5.7 1.1 0.6 μg/mL Error 0.01 19 100.8 15.6 4.7 0.6 0.2 0.2

[0373] A graph on mean drug concentration values is presented in FIG.24, showing a GSSG Pt pharmacokinetic curve for intravenousintroduction. TABLE D Pharmacokinetic parameters Estimation on Directmethod Error model Error D, mg/kg 10 10 Dose Cmax, 187.0 19 186.4 18Maximal concentration μg/mL m, min 1 1 1 1 Time for maximum attainmentAUC, 60 min- 1800 165 1790 178 Square under the curve before μg/mL 60min. AUC t min- 1825.0 172.7 1797.5 171 Full square under the curveμg/mL AUCt/Cmax, 9.8 0.9 9.65 0.85 Effective duration min.

Conclusion

[0374] It was demonstrated at the comparative analysis for the GSSG.PTand GSSG drugs that the main GSSG.Pt pharmacokinetic parameters (maximalblood concentration and effective drug influence duration) obtained bothby the direct method and by the estimation on model exceed about 2-3times the main GSSG pharmacokinetic parameters. The integral indicesshowing the drug blood presence duration determined by calculation ofthe square under the GSSG.Pt pharmacokinetic curve exceed 4 times thecorresponding indices regarding to GSSG. Thus, due to more advantageouspharmacokinetic parameters the GSSG.Pt drug has higher pharmacologicalactivity and biological availability comparing to the GSSG drug.

EXAMPLE 5 Effect of GSSG.Pt and GSSG on Cytokine Production by HumanPeripheral Blood Mononuclear Leukocytes in Vitro

[0375] Oxidized glutathione (GSSG) as well as a structural analogthereof, which is a hexapeptide with a stabilized disulfide bond, wereevaluated for their effect on cytokine production by human peripheralblood mononuclear leukocytes in vitro.

[0376] The leukocytic cytokine production was triggered by adding amitogen, concanavalin A (ConA) to the cell culture immediately afterintroducing the test substances. In 24 hours of the cellular exposure toConA and the test articles, the culture supernatants were sampled andstored until cytokine determination at −70° C.

[0377] With the aim of evaluating the functional status of the cells andtheir capacity of responding to the mitogen in the presence of the testarticles at each concentration level, the control cell cultures,containing the test articles in identical concentrations, were incubatedfor 72 hours following the initial concomitant introduction of ConA andthe test substances. Sixteen hours prior to the incubation completion,³H-thymidine was added, and the label rate of incorporation into DNA wasinterpreted as the criterion of the cellular test system functionalstate.

[0378] Venous blood from male healthy volunteers was collected intoplastic heparinized tubes (endotoxin tested). PMNL fraction was isolatedby centrifugation in density gradient of Ficoll and sodium diatrizoate(Histopaque-1077; Sigma).

[0379] Cell concentration was adjusted to 2×10⁶ per 1 mL of culturemedium (RPMI 1640, Sigma) containing: HEPES (20 mM); L-glutamine (2 mM);Gentamicin (50 mg/mL); fetal calf serum (10%). All the reagents usedwere of cell culture tested grade, Sigma. Cell viability was estimatedby the Trypan blue exclusion method and 100 mL of cell suspension(200,000 cells) were placed into each well of flat bottom 96-wellsterile microtiter plates for tissue cultures. Cells from each subjectwere placed into no less than 39 wells.

[0380] The five following final concentrations of the test articles(GSSG, as well as GSSG.Pt) were evaluated: 5000 mg/mL; 500 mg/mL; 50mg/mL; 5 mg/mL; and 0.5 mg/mL. Each concentration was established in noless than six wells by adding 50 mL of medium containing the appropriatequantity of the previously dissolved test articles. Another six wellswere used for control cultures: only 50 mL of medium was added.

[0381] Immediately after the test articles had been introduced into thecultures, 50 mL of medium containing ConA (Sigma, cell culture tested)in a quantity required for a final concentration of 4.0 mg/mL, was addedto all the wells excepting three additional ones which served forevaluation of spontaneous ³H-thymidine uptake (without ConA).

[0382] After a 24-hour incubation at 37° C. and 5% of CO₂, contents ofthree wells (from each sextuplet of identical wells) were taken out,centrifuged, and the supernatants were frozen and kept at −70° C. untilthe cytokine assay was to be performed. Cultures in the other threewells (of each sextuplet) were incubated further under the conditionsdescribed above.

[0383] Fifty-six hours after the incubation had begun, 1.0 mCi of³H-thymidine was added into all the remaining cultures, the plates wereincubated for another 16 hours, and then the contents of the wells wereharvested and transferred onto glass-fiber filters which wereconsequently treated with 5% trichloroacetic acid and ethanol. Thefilters were dried and their radioactivity (counts per minute, cpm) wasdetermined using liquid scintillation counter, Betaplate 1205 (LKB).

[0384] Mean radioactivity values for triplicates of identical cultureswere used to calculate the index of mitogenic stimulation: the ratio ofaveraged cpm values for ConA stimulated cultures to averaged cpm valuesfor unstimulated ones (three wells without ConA). This stimulation indexfor wells, where the test articles were present in variousconcentrations, served as a criterion of cellular functional status, andability of the cells to respond to mitogenic stimulation.

[0385] Supernatants of 24-hour culture triplicates were subsequentlyassayed for cytokine content only if their 72-hour matched controlculture triplicates developed mitogenic response to ConA with value ofthe stimulation index in the range from 15 to 50.

[0386] Concentrations of interleukin-1b (IL-1b), interleukin-2 (IL-2);interleukin-3 (IL-3); interleukin-4 (IL-44); interleukin-6 (IL-6),interleukin-8 (IL-8); interleukin-10 (IL-10); interleukin-12 (IL-12);tumor necrosis factor-α, g (TNF-(α, g), and interferon-α, g (IFN-α, g)were determined by ELISA using commercial reagent kits (Medgenix,Belgium) and were expressed in pg/mL of culture supernatants.

[0387] The salient findings given in Tables 8, 9. As one can see fromTables 8 and 9, the adding of GSSG and GSSG.Pt into the culture mediaresulted in statistically significant and dose-dependent stimulation ofthe cytokine production by human mononuclear leukocytes. However,GSSG.Pt stimulating influence was more significant (1.5-2 times as much)on the studied cytokine production with stimulation and regulation forproduction of the wider cytokine range in comparison with the GSSGeffect. One can clearly see correlation of the interrelated cytokinechanges (increasing of IL-1b, IL-2, TNF-α, g along with decreasing ofIL-4, IL-10) in the Tables 8 and 9.

[0388] Thus, the GSSG.Pt impact on the human peripheral mononuclearleukocytes in vitro was manifested with considerable stimulation of thewider cytokine range release into culture media considering theirreciprocal regulative effect, and, thereby, it confirmed the GSSG.Ptstimulatory and regulatory effect on the natural cytokine-producingcapacity of the human blood cells.

EXAMPLE 6 Effect of GSSG and GSSG.Pt on Cytokine and Hemopoietic FactorProduction as Well as on Hemopoiesis and Immunity Parameters inCyclophosphamide-Induced Hemo- and Immunodepression

[0389] 1. The oxidized (GSSG) glutathione as well as the structuralanalog thereof, which is the hexapeptide with the stabilized disulfidebond, were evaluated in a murine model of hemo- and immunodepressioninduced by a single administration of cytostatic agent Cyclophosphamide(CP).

[0390] The study was designed to evaluate the effect of a five-day longadministration of the test articles on the capability of the CP-treatedmurine splenocytes to produce interleukin-1 (IL-1 α,b); interleukin-2(IL-2); interleukin-3 (IL-3); interleukin-4 (IL-4); interleukin-6(IL-6), interleukin-8 (IL-8); interleukin-10 (IL-10); interleukin-12(IL-12); tumor necrosis factor-α, g (TNF-α, g), interferon-α, g (IFN-α,g) and G-CSF, M-CSF, GM-CSF in vitro. In addition, the blood leukocyteand lymphocyte count and the bone marrow cellularity (karyocyte count)were determined at eight days after CP administration. Some animalsreceiving CP were then challenged with sheep red blood cells (SRBC), andthe humoral immune response to the antigen was evaluated.

[0391] Male CBA mice (180 to 200 g body weight) were given a singleintroperitoneal injection of CP in a dose of 50 mg/kg. Four groups ofanimals (with no less than 15 mice in each) were formed. The groupdescription is represented below.

[0392] Control groups:

[0393] #1—intact animals receiving a single injection of normal saline(NS) instead of CP injection, which further were treated with testarticle vehicle (normal saline);

[0394] #2—animals receiving a single CP injection, which further weretreated with test article vehicle (normal saline);

[0395] Test groups:

[0396] #3—animals receiving a single CP injection, which further weretreated with the test article (GSSG dissolved in normal saline) in adose of 5 mg/kg;

[0397] #4—animals receiving a single CP injection, which further weretreated with a GSSG.Pt (dissolved in normal saline) in a dose of 5mg/kg.

[0398] Twenty-four hours after the CP injection, five animals in eachgroup were immunized with SRBC (10⁷ cells in 0.5 mL of NS,intra-peritoncally).

[0399] On day 3 after the CP injection (24 hours after the immunization)the intraperitoneal injections of the test or reference articles werestarted (as it has been described above). Injections were performedduring five days: once a day, daily.

[0400] Twenty-four hours after the completion of five-day treatmentcourse (on the 8^(th) day after the CP injection), mice were euthanizedand splenocyte cultures were aseptically prepared for assessment ofspontaneous cytokine and hemopoietic factor production by the spleenlymphocytes in vitro.

[0401] Simultaneously, blood and marrow samples were collected for bloodleukocyte, lymphocyte, and marrow nucleated cell counts.

[0402] Serum samples from immunized animals were tested on level of SRBCagglutilnins (day 8 after the CP injection, and day 7 after theimmunization).

[0403] Table 10 shows the parameters of cytokine and hemopoietic factorproduction by splenocytes, bone marrow and blood count indices, and theimmune response to sheep red blood cells in mice receiving the testarticles against the background of cyclophosphamide induced hemo- andimmunodepression.

[0404] According to the Table 10 data, the GSSG.Pt administration set tonorm the cytokine and hemopoietic factor production while GSSG performedonly exiguous stimulating effect. Besides, GSSG.Pt stimulated productionfor the wider cytokine and hemopoietic factor range as well as hadsignificant regulatory influence on shift with respect of the cytokinestatus that was confirmed with the positive correlation of theinterrelated cytokine changes at the corresponding pathologic process.

[0405] Thus, the GSSG.Pt use in CP-induced hemo- and immunocompromisedanimals results in a prominent stimulation and regulation on thecytokine and hemopoietic factor endogenous production along withrestoration of the bone marrow and blood cellular indices as well asimmune response development to sheep red blood cells.

[0406] 2. The purpose of the present study was to explore the GSSG_Ptefficacy on the cyclophosphamide-induced cytopenia (myelopenia) model.

[0407] The study was conducted on the white male rats weighing 160.0 gr.Cyclophosphamide was introduced once subcutaneously at the back in doses50 mg/kg (a vehicle was water for injections).

[0408] Four groups of animals (with no less than 15 mice in each) wereformed. Group description is represented below.

[0409] Control groups:

[0410] #1—intact animals receiving a single injection of normal saline(NS) instead of CP injection, which further were treated with testarticle vehicle (normal saline);

[0411] #2—animals receiving a single CP injection, which further weretreated with test article vehicle (normal saline).

[0412] Test group:

[0413] #3—animals receiving a single CP injection, which further weretreated with GSSG.Pt (dissolved in normal saline) in a dose of 5 mg/kg.

[0414] On day 2 after the CP injection the intraperitoneal injections ofthe test or reference articles were started (once a day, during 10-15days).

[0415] At the end of each series (10 and 15 days) the experimentalgroups were euthanized through ether overdose and peripheral blood (tailveins) and bone marrow (femora) were taken for the analysis. Thehematological studies were conducted with the unitized standard methods.The peripheral blood analysis results are represented in the Table 11;myelogram analysis results are given in the Table 12.

[0416] Analyzing Table 11 results one can note that cyclophosphamide inthe dose 50 mg/kg was found to perform the marked cytopenic effect atall of the formed blood elements with absolute and relative lymphopeniadepending on the observation terms (maximally exhibited at the day 15).

[0417] It should be noted that four animals died: on the 9^(th),10^(th), 12_(th), and 14^(th) days. In fact, all these animalsdemonstrated flabbiness, hypodynamia, weight loss.

[0418] The GSSG.Pt introduction provided the significant stimulatingeffect. The general state improvement, positive weight changes wereobserved, and the immature cell form appearing in blood indicated thebone-marrow hemopoiesis activation. The death of the animals was notnoted.

[0419] Analysis of the myelogram revealed that cyclophosphamide in thedose 50 mg/kg was found to perform the significant myelotoxic effect.Erythro-, trombocyto- and lymphopoiesis are especially suppressed.Myelosuppression is the most marked at the 15^(th) day.

[0420] The GSSG.Pt administration had the considerable myelostimulatingeffect.

[0421] The drug GSSG.Pt in the dose 15 mg/kg being introduced as thedaily course during 10-15 days exerts the marked myelostimulating effectat the cyclophosphamide-induced cyto- and myelopenia. The drugadministration improves the general state, it positively influences atthe body weight changes and lowers mortality by about 30%.

EXAMPLE 7 Effect of GSSG and (Li.GSSG.Pt) on Cytokine and HemopoieticFactor Production as well as on Hemopoiesis and Immunity Parameters inRadiation-Induced Hemo- and Immunodepression

[0422] Both oxidized (GSSG) and the structural analog thereof, which isthe lithium salt of the hexapeptide with the stabilized disulfide bond(Li.GSSG.Pt), were evaluated in a murine model of hemo- andimmunodepression induced by a single irradiation in a total dose of 1Gy.

[0423] The study was designed to evaluate efficacy of seven-day dailyadministration of the test articles (with the dosing started two hourspost-exposure) on the capability of the splenocytes from mice exposed toradiation to produce interleukin-1 (IL-1 α,b); interleukin-2 (IL-2);interleukin-3 (IL-3); interleukin-4 (IL-4); interleukin-6 (IL-6),interleukin-8 (IL-8); interleukin-10 (IL-10); interleukin-12 (IL-12);tumor necrosis factor-α, g (TNF-α, g), interferon-α, g (IFN-α,) andG-CSF, M-CSF, GM-CSF in vitro. In addition, the blood leukocyte andlymphocyte counts and the spleen and bone marrow cellularity (karyocytecount), as we well as splenic and medullary colony-stimulating capacity,were determined at the 8^(th) day post-exposure.

[0424] Male CBA mice (18 to 20 g body weight) were irradiated withsingle dose of 180 kV X-rays filtered with 0.5 mm Cu (at 15 mA,distance—70 cm, duration two min. and 28 sec.). The total absorbed dosecomprised approximately 1 Gy.

[0425] Four groups of animals (with no less than 12 mice in each) wereformed. Group description is represented below.

[0426] Control groups:

[0427] #1—intact animals receiving a sham irradiation procedure toreproduce a stress impact, which further were treated with the testarticle vehicle (normal saline);

[0428] #2—control animals irradiated in a dose of 1 Gy, which furtherwere treated with test article vehicle (normal saline).

[0429] Test groups:

[0430] #3—animals irradiated in a dose of 1 Gy, which further weretreated with the test article (GSSG dissolved in normal saline) in adose of 5 mg/kg;

[0431] #4—animals irradiated in a dose of 1 Gy, which further weretreated with the test article (Li.GSSG.Pt dissolved in normal saline) ina dose of 5 mg/kg.

[0432] Two hours after the irradiation the intrapcritoneal injections ofthe test or reference articles were started (as it has been describedabove). Injections were performed during seven days: once a day, daily.

[0433] Twenty-four hours after the completion of seven-day treatmentcourse (on the 8^(th) day after the irradiation), mice were euthanizedand splenocyte cultures were aseptically prepared for assessment ofspontaneous cytokine and hemopoietic factor production (interleukin- 1(IL-1 α,b); interleukin-2 (IL-2); interleukin-3 (IL-3); interleukin-4(IL-4); interleukin-6 (IL-6), interleukin-8 (IL-8); interleukin-10(IL-10); interleukin-12 (IL-12); tumor necrosis factor-α, g (TNF-α, g),interferon-α, g (IFN-α, g) and G-CSF, M-CSF, GM-CSF) by the spleenlymphocytes in vitro.

[0434] Simultaneously, blood, spleen and marrow samples were collectedfor blood leukocyte and lymphocyte, and spleen and marrow nucleated cellcounting.

[0435] Additionally, hemopoietic colony-formation ability of spleen andbone marrow cells was assessed by the method of colony-forming unit(CFU) direct count in the spleens of irradiated singenic CBA micereceiving intravenously spleen or bone marrow cells obtained fromanimals of control or test groups.

[0436] Splenocytic levels for the cytokine and hemopoietic factorproduction, blood, bone marrow, and spleen cellular indices as well asthe colony-forming parameters (colony-forming units, CFU) at the bonemarrow and spleen of the irradiated animals at the 8^(th) daypost-exposure, are summarized in Table 13.

[0437] As is evident from the table data, the Li.GSSG.Pt administrationresults in statistically significant recovery of the cytokine andhemopoietic factor production by splenocytes, whereas GSSG produces lesssignificant effect. However, Li.GSSG.Pt influences endogenous productionfor the wider cytokine and hemopoietic factor range as well as regulatesthe cytokine status alterations with respect of the correspondingpathologic process.

[0438] Thus, the Li.GSSG.Pt usage as an applied method in animals withdeveloped radiation-induced hemo- and immunodepression results inpronounced stimulation-regulation of the endogenous cytokine andhemopoietic factor production, and also leads to an effective recoveryfor the cellular compositions of the blood, lymphoid and hemopoieticorgans as well as the bone marrow and spleen colony-forming activity.

EXAMPLE 8 Effect of the GSSG.Pt Composite and Salts thereof on Processesof Phosphate Modification as Well as on Content of Lymphocytes BearingIL-2 Receptors

[0439] The molecular mechanisms of the reproduction of theimmuno-biochemical effects of the cytokines with the GSSG.Pt compositeand the salts thereof were studied.

[0440] At the study action of the GSSG.Pt composite and the saltsthereof—sodium (Na), lithium (Li) and magnesium (Mg)—was evaluated atthe murine model of hemo- and immunodepression induced by singleadministration cytostatic cyclophosphamide (CF).

[0441] At this study the effect of a five-day long administration of thetest articles on the capability of the phosphorylating level of thelymphocyte cytosol proteins on tyrosine and content of the “active”lymphocytes-carriers of IL-2-receptors were evaluated.

[0442] Male CBA mice (18 to 20 g body weight) were given a singleintraperitoneal injection of CP in a dose of 50 mg/kg. After the CFinjection the animals were introduced with the tested articles in doseof 5 mg/kg 24 hours later. The tested articles were introduced duringfive days (daily, once a day). Twenty-four hours after completion of thetested article introduction the mice were euthanized and blood sampleswere collected to conduct the study.

[0443] A fraction of mononuclear leukocyte was obtained bycentrifugation in gradient of ficoll-metrizoat (Histopaque, Sigma). Cellconcentration was adjusted to 2×10⁶ cells per 1 ml of cell culturemedium (RPMI 1640), containing 20 mM HEPES, 2 mM glutamine, 50 mg/mLgentamicin and 10% fetal calf. Cell viability was estimated by theTrypan blue exclusion method, then the cell suspension was placed intowells of 96-well microliter plates —200,000 cells per well.

[0444] Content of the lymphocytes-carriers of the IL-2-receptors wasdetermined according to Horgan A. F. (1994) on smears of mononuclearslip. Mononuclear antibodies to chains p55 and p75 of the IL-2-receptorwere used as the first antibodies. To reveal the first antibodies thepolyclonal rabbit antibodies against murine immunoglobulins marked withwere used. Count of the lymphocytes-carriers of the IL-2-receptors wasmade in percentage to the number of total lymphocytes.

[0445] For metabolical marking lymphocytes were cultivated in the Iglamedium with addition of 10% cattle serum. The metabolical marking with[32P]ortho-phosphoric acid was performed by cell incubation during 10-12hrs. in the phosphorusless medium DME containing 100 μCi/ml of[32P]ortho-phosphate. On each sample 0.2 ml of the medium with isotopewere added. After incubation cells were destroyed by pipetting andcentrifuged at 6000 g for 30 min. The obtained supernatant was used forimmunoprecipitation with polyclonal antibodies to phosphotyrosinie at Fumethod (1992). Protein A-sefarose was used for the precipitation ofimmune complex. The precipitate was washed three times and theprecipitate activity was counted on “Gamma” counter.

[0446] According to the results of the conducted study (Table 14), itwas found that action of the GSSG.Pt composite on the isolatedlymphocytes causes (see Table 14) at 10 minute a significant increase ofthe phosphorylating level on tyrosine of the lymphocyte cytosolproteins, which is the integral indication of the activity of thesignal-transducing systems. These changes due to the GSSG.Pt compositeaction, largely due to the GSSG.Pt composite derivative action determinethe modulation of the redox-sensitive gene expression, first of all,immunologically important genes, responsible for the synthesis of thecytokines and hemopoietic factors.

[0447] In Table 15 the data is given on phosphorylating level ontyrosine of the lymphocyte cytosol proteins and percentage of thelymphocytes-carriers of IL-2-receptors in CBA-line mice received thetested articles having cyclophosphamide induced hemo- andimmunodepression.

[0448] Application of the GSSG.Pt composite salts results in theincrease of the percentage of the lymphocytes-carriers of IL-2-receptorsand almost normalizing their quantity (normal one is 18.3±1.6%). Thesimilar regularity was found when phosphorylating level on tyrosine ofthe lymphocyte cytosol proteins was studied.

[0449] Application of the GSSG.Pt composite salts results in restorationof the percentage of the lymphocytes-carriers of IL-2-receptors inimmunodeficiency conditions modeled by cyclophosphamide. There is theincrease of the phosphorylating level on tyrosine of the lymphocytecytosol proteins of the signal-transducing systems that can be one ofthe factors of the described immunostimulating actions of the articlestested.

[0450] Thus, the example provides evidence of GSSG.Pt to reproduce(imitate) the regulatory effects of the range of cytokines, first ofall, IL-2. We are intending to mean the induction by the GSSG.Ptcomposite of the intracellular mechanisms performing regulatory cytokinesignals on the system of immunocompetent and hemopoietic cells as theIL-2 effect reproduction. The conducted studies have shown that changingof the phosphorylating level on tyrosine of the lymphocyte cytosolproteins of the signal-transducing systems of the cells of the organsimmunogenesis and hemopoiesis in conditions of cyclophosphamide modeledimmunodeficiency causes the effect of the dynamic normalization of theactive lymphocyte content.

[0451] Therefore, application of the GSSG.Pt composite and derivativesthereof in the form of therapeutically purposed medicinal drugs not onlystimulates the cytokine and hemopoietic factor endogenous production butalso provides the reproduction of the immune-biochemical cytokineeffects, especially in case of receptors desensitization observed mainlyin oncological and retro-virus pathology.

EXAMPLE 9 Stimulation of Endogenous Cytokine Production and theTherapeutic Effect of the GSSG_Pt Application in a Patient with aStomach Cancer, Peritoneal Metastases, Ascites, Splenomegaly andCholestatic Hepatitis

[0452] A 33-year old patient was diagnosed as having stomach neoplasmfor more than two years (adenocarcinoma of moderate differentiationdegree). In 1993 the patient was operated for malignant stomach ulcerand numerous dense lymph nodes were found in the porte hepatis whichwere considered to be metastases.

[0453] In January 1994 the course of chemotherapy (5-FU) was complicatedby the severe cholestasis and percutaneous drainage of the left andright liver ducts was undertaken, that six months later was followed bythe choledochoejunostomy with changeable transliver drains with Brown'sanastomosis.

[0454] In November 1995 the patient's state worsened. According to theexaminations the patient experienced an active secondary hepatitis. Theliver was enlarged and painful and protruded from the costal arch up to5-6 cm. Blood chemistry indices proved to be persistently abnormal andhardly corrected by the performed treatment: bilirubin—40.0 due toindirect (up to 31.0); activity of amino-transferases - approximatelysix times higher than upper normal limit, hypoalbunemia was up to 26%;and there was also hypergammaglobulinemia; hypercholesterolemia was upto 10.2 mmol/l.

[0455] During fibrogastrocopy (November, 1995) a stomach cancer locatedin the middle area of the stomach body and extended about 8 cm wasconfirmed. The tumor was solid-like type. Stomach walls were rigid.Histology examination defined the tumor as adenocarcinoma of moderatedegree differentiation. In December, 1995, the patient had anexplorative laparotomy. Ascites was found with plural metastases allover the peritoneum, splenomegaly. The patient's case was identified asinoperable.

[0456] A decision was taken to apply GSSG drug form. The drug wasinjected parenterally (intramuscularly and intravenously), andadditionally, the drug form was used via local injections around thetumor tissue through an endoscope. An average doses which were used forintramuscular and intravenous injections—0.1-0.5 mg/kg, and for localinjections—up to 50 mg in situ. Parenteral injections of the drug wereapplied every other day, b.i.d. (intravenous injections at the morning,and intramuscular ones at the evening), during three weeks, and afterthat, two times a week during four weeks. The drug introduction throughthe endoscope was performed once in seven days. Two months after thebeginning of the treatment with the drug form used thefibrogastroduodenoscopy showed: esophagus was passable, mucous membranewas pink, cardia rosette was partly closed. On empty stomach moderateamount of foamy secretion was in the stomach, which was intensivelycolored with bile. The tumor extent was 4.8 cm. At the same time,substantial improvement of hematology and blood chemistry indices wasfound and the liver size decreased up to 3 cm (below the costal arch).

[0457] Six months after the treatment completion (July, 1996) thepatient's state worsened significantly. According to the examination thesecondary hepatitis relapsed. The liver was increased in size andtender, protruded 4 cm beyond the costal arch. Blood chemistry indiceswere abnormal and hardly corrected by the performed treatment:bilirubin—360 due to indirect (up to 28.0); activity ofamino-transferases—approximately four times higher than upper normallimit, hypoalbunemia was up to 21%; and there was alsohypergamnmaglobulinemia; hypercholesterolemia was up to 9.42 mmol/l.

[0458] At the fibrogastrocopy (July, 1996) a stomach cancer located inthe middle area of the stomach body and extended about 6 cm wasconfirmed. The tumor was solid-like type. Stomach walls were rigid.Histology examination defined the tumor as a moderate degreedifferentiation adenocarcinoma. In August, 1996, the patient had anexplorative laparotomy. Ascites was found with plural metastases allover the peritoneum, splenomegaly. Considering the previously conductedtherapy the decision was taken to apply the new drug GSSG.Pt that is thestructural analog of the former administered GSSG drug. The drug wasintroduced according to the identical regime: parenterally(intramuscularly and intravenously), and additionally, the drug form wasintroduced via local injections around the tumor tissue through anendoscope. An average doses which were used for intramuscular andintravenous injections—0.1-0.5 mg/kg, and for local injections—up to 50mg in situ. Parenteral injections of the drug were applied every otherday, b.i.d. (intravenous injections at the morning, and intramuscularones at the evening), during three weeks, and after that, two times in aweek during four weeks. The drug introduction through the endoscope wasperformed once in seven days.

[0459] Two months after the treatment initiation with the applied drug:the liver protruded 1 cm beyond the costal arch, tenderless atpalpation. According to the ultrasound examination data: there isfibrous tissue at the place of previously determined cancer sites. Atthe fibrogastrocopy: the esophagus was passable, mucous membrane waspink, cardia rosette was partly closed. The gastric walls are elastic.There was moderate amount of foamy secretion in the stomach with salivain empty stomach. The tumor extent was 1.5 cm. The duodenum was freelypassable. At the same time, substantial improvement of hematology andblood chemistry indices was indicated.

[0460] Comparing the therapeutical efficacy of the drugs GSSG.Pt andGSSG using of the former one was found to be advantageous that wasmanifested by the positive changes at the clinical, biochemical,hematological and immunological indices, fibrogastrocopy data (the tumorsize decrease at 75% while applying GSSG.Pt comparing to the 40%decrease after the GSSG administration) (Tables 16, 17). Moreover, atthe Table 17 one can see that GSSG.Pt stimulates production of the widercytokine and hemopoietic factor range having a regulatory influence ontheir content change.

[0461] Thus, the treatment according to the present invention resultedin considerable regress of tumor process with simultaneous obviousbeneficial changes in hematology, blood chemistry and immunologyparameters, and significant improvement of the life quality.

EXAMPLE 10 Therapeutical Efficacy the GSSG.Pt Application for Treatmentof Lung Cancer

[0462] No. 1

[0463] Year of birth: 1938.

[0464] Diagnosis: Cancer of the right lung upper lobe.

[0465] Histological diagnosis: No. 45760 (State Research Center onPulmonology)—small cell cancer.

[0466] Case-history: No. 4024.

[0467] Complaints on admission: coughing with hard discharged mucoussputum, dyspnea on little exertion.

[0468] Objective examination: The patient's state is satisfactory.Peripheral lymph nodes are not enlarged. There are coarse breath soundsweakened at the upper and medium regions of the right lung. There arerare dry rales, dyspnea on the slightest exertion.

[0469] Roentgenography (initial data): The upper mediastinum shadow isbroadened due to enlarged right paratracheal lymph nodes. The indistinctshadow of the upper right root part is broadened. There is an additionalshadow in the peripheral S₃ region of the right lung against thebackground of marked interstitial changes in both lungs. The rightinterlobar borders are thickened. Conclusion: there are signs ofmetastases into lymph nodes of the root and the mediastinum withlymphangoitis sings.

[0470] Treatment course: there were applied three immunochemotherapycourses using GSSG.Pt drug.

[0471] After the treatment: the patient's state has improvedsignificantly: the mild weakness is still present, there is no dyspnea.

[0472] Objective examination: The state is satisfactory. The peripherallymph nodes are not enlarged. There is a weakened breath sounds in themedium departments of the right lung. The breath sounds in otherdepartments are vesicular. There are no rates.

[0473] Roentgenography (after the treatment performed): The lung fieldsare particularly clear. There are mild infiltrative signs at the S₃department of the right lung. The roots are not enlarged. There are noadditional formations at the right root projections. The uppermediastinum shadow is not enlarged. The solitary paratracheal lymphnodes can be determined.

[0474] No. 2

[0475] Year of birth: 1945.

[0476] Diagnosis: Right lung cancer, hepatic metastases.

[0477] Histologic diagnosis: No. 45998, small cell cancer

[0478] Case-history: No. 4076

[0479] Complaints on admission: coughing with hard discharged mucoussputum, dyspnea on mild exertion, weakness, constant pains in the lumbarregion extending to stomach. During last six months the patient lost 6kg.

[0480] Objective examination: The state is medium severe. Scleras areicteric. The breathing sounds are coarse, weakened in the right lungupper and medium departments. In the right supraclavicular zone one canpalpate enlarged lymph nodes (solid consistency, hardly movable,size—3.0 and 1.0 cm, painless). At auscultation there are coarsebreathing sounds, weakened in the medium departments in the right. Thereare solitary dry rales, dyspnea on the slightest exertion. The liverprotruded 3.5 cm over the costal arch.

[0481] At examination there were found: middle-lobe bronchus cancer,middle lobe hypoventilation, pneumonitis. At ultrasound examinationthere are sings of metastatic liver impairment.

[0482] Roentgenography (initial data): The right lung middle lobe isdecreased in size (state of hypoventilation). Against the background ofthe increased lung pattern there is intensive, almost homogenousinfiltration with distinct margin along the horizontal interlobarpleura. The right root cannot be determined. The upper and lower lobesof the right lung and the left lung are without any particular features.The mediastinal organs are not noticeably displaced.

[0483] Treatment course: there were applied three immunochemotherapycourses using GSSG.Pt drug.

[0484] After the treatment: The patient's state has improvedsignificantly: there are no weakness and dyspnea, appetite has appeared,he gained 5 kg. The blood indices have restored.

[0485] Objective examination: The state is satisfactory. The peripherallymph nodes are not enlarged. There is a weakened breath sounds in themedium departments of the right lung. The breath sounds in otherdepartments are vesicular. There are no rales.

[0486] According to the liver ultrasonography and CAT scan data: thereis full mass process regression.

[0487] Roentgenography (after the treatment performed): There is theinsignificant atelectasis of the middle lobe at the thoracic X-rayspicture. The roots are structural and not enlarged, the right one isslightly displaced downward. The heart is not enlarged in size.

[0488] Comparing to the initial data there is considerable positivedevelopment: at the right the pulmonary tissue has become moretransparent (reduction of the hypoventilation signs), there are noinfiltrative shadows.

Conclusion

[0489] Evaluating therapeutical effectiveness of the GSSG.Pt drug thefollowing was found:

[0490] 1. Clinical restoration of indices (general state improvement,pathologic absence of symptoms, body weight gaining);

[0491] 2. Roentgenologic picture changes (pulmonary tissue transparencyincrease, infiltrative absence of shadows, disappearing of atelectasisand hypoventilation);

[0492] 3. Hematologic restoration of indices(increase of the erythrocytecount and hemoglobin content, restoration of white blood count);

[0493] 4. Changes of the ultrasound and CAT scan data (full tumorprocess regression);

[0494] 5. Immune indices restoration and increase of the CD16⁺, CD25+counts indicating restitution of the antitumor surveillance system.

[0495] 6. Induction of the wide cytokine range synthesis as well asmodulation of their content mutual regulation (correlation of thecontent changes of IL-1b, IL-4, TNF-α).

[0496] Thus, in the given clinical Examples it has been demonstratedthat the GSSG.Pt drug application has provided faster restoration ofclinical, roentgenologic, hematologic and immune indices ensuring moreeffective restitution of the immunity and hemopoiesis systems. Theaforementioned indicates the tumor process regression that, eventually,calls out significant increase of the patient's quality of life.

EXAMPLE 11 Therapeutical Efficacy the GSSG.Pt Application for Treatmentof Chronic Viral Hepatitis B (CHBV)

[0497] No. 3

[0498] Date of birth: 1945

[0499] Diagnosis: CHBV, replicative phase (PCR HBV+) with moderateactivity grade.

[0500] Case-history: No. 1068.

[0501] Complaints at admission: weakness, discomfort under the rightcostal arch, nausea, no appetite.

[0502] Anamnesis morbi: during last six years the patient notedperiodically appeared dull pains under the right costal arch, weakness,urine color changes. At examination there was found: hyperbilirubinemiaup to 34 mmol/L, ALT increase to 5.4 mmol/hr.L. CHBV serologic markersand PCR HBV(+) were determined at the hospital examination.

[0503] Objective examination: The patient's state is satisfactory. Theliver protruded 2 cm beyond the costal arch. The liver margin is firmand tender.

[0504] Previous treatment was not performed.

[0505] Treatment course: There was performed the treatment course withadministration of the GSSG drugs according to the regime.

[0506] State after the performed treatment course: There were noted thefollowing positive changes—significant general state improvement, noweakness and nausea, diminution of the to discomfort sensation. Theliver protruded 0.5 cm beyond the costal arch. The liver margin is softand tenderless.

[0507] No. 4

[0508] Year of birth: 1964

[0509] Diagnosis: CHBV, replicative phase (PCR HBV+), moderate activitydegree.

[0510] Case-history: No. 1043.

[0511] Complaints on admission: considerable weakness, no appetite,sweating, urine darkening.

[0512] Anamnesis morbi: The patient feels sick beginning from January1996 when for the first time there appeared dull pains under the rightcostal arch, temperature increase up to 38.7° C., vomiting, urinedarkening. Acute viral hepatitis B was diagnosed and confirmedserologically. The patient was administered with detoxicating andantibacterial therapy. However, afterwards there were observed increasedHbs Ag and ALT values, persistent viral replicative activity. Beingexamined at the last time there were found: hyperbilirubinemia to 78mmol/L, ALT increase to 6.2 mmol/hr.L. CHBV serological markers and PCRHBV(+) were determined at the hospital examination.

[0513] Objective examination: The general state is satisfactory. Skinand scleras are icteric. The liver protruded 2.5 cm beyond the costalarch. The liver margin is firm and tender.

[0514] Previous treatment: from 17.09.97 the patient was administeredwith acyclovir during 21 days. After the course completion there wereincreased ALT—up to 2.1 mmol/hr.L, bilirubin to 32 mmol/L. The Hbs Agdegree did not change. The patient's state was defined by thesignificant asthenic-vegetative syndrome.

[0515] Treatment course: There was performed the treatment course withadministration of the GSSG.Pt drugs according to the regime.

[0516] State after the performed treatment course: There were noted thevivid following positive changes—significant general state improvement,no weakness, sweating and nausea. The skin and scleras were not icteric.The urine color became normal, diminution of the discomfort sensation.The liver protruded 0.5 cm beyond the costal arch. The liver marginbecame softer and tenderless.

[0517] Comparison (Tables 20, 21):

[0518] At the comparative analysis on the therapeutical efficacy of theGSSG.Pt and GSSG drugs the former was shown to be advantageous that wasmanifested by the following:

[0519] 1. Biochemical indices normalization (ALT and bilirubindecrease);

[0520] 2. Significant HBs Ag decrease and replication termination;

[0521] 3. Immune indices normalization and increase of the CD95+ contentindicating the apoptosis process activation in the virus-transformedcells;

[0522] 4. Considerable regulation for the wider cytokine range.

EXAMPLE 12 Therapeutical Efficacy the GSSG.Pt Application for Treatmentof Acute Viral Hepatitis B (AHBV)

[0523] No. 5

[0524] Sex: female.

[0525] Age: 20.

[0526] In-patient card: 678

[0527] Diagnosis: Acute viral hepatitis B (HBs Ag “+”), replicativephase (PCR HBV “+”), prolonged form; chronic viral hepatitis D,replicative phase (PCR HDV “+”).

[0528] Complaints during examination: weakness, appetite decrease.

[0529] Anamnesis morbi: The patient felt sick in August 1997, when shenoticed sharp weakness, malaise, back bone aching, temperature raisingup to 38.8° C. Dark urine and sclera icterus appeared 10 days later. Thepatient was admitted to the viral infectious clinic, where she receivedcourse of the detoxicating, spasmolytic, antibacterial therapy. However,replicative viral activity and increased GPT level were still present.The prolonged cytolytic syndrome gave foundation for administration ofthe drug GSSG.Pt.

[0530] Previous treatment was not performed.

[0531] Treatment with GSSG.Pt drugs: from 30.10.97 to 23.11.

[0532] Patient's state after the treatment course completion:

[0533] The patient's state is satisfactory. She noted the appetiteincrease, weakness reduction.

[0534] Conclusion (Tables 22-24):

[0535] The immunomodulating course with the GSSG_Pt drugs has providedthe following positive changes: biochemical indices normalization;termination of HBV and HDV replication; termination of HBs Agpersistency; virus-infected cell apoptosis induction; general stateimprovement; stable therapeutic effect.

EXAMPLE 13 Therapeutical Efficacy the GSSG.Pt Application for Treatmentof Chronic Viral Hepatitis C (CHCV)

[0536] No. 6

[0537] Sex: male.

[0538] Age: 18.

[0539] Patient's case: No. 1043

[0540] Diagnosis: Chronic viral hepatitis C, replicative phase (PCR HCV“+”), moderately manifested activity; chronic viral hepatitis B,integrative phase (PCR HBV “−”); narcotic intoxication, narcomania.

[0541] Complaints during examination: weakness, pains at the right underthe ribs, at knee-joints, the backbone and wrist joints.

[0542] Anamnesis morbi: The patient noticed pains in the knee-joints andthe backbone at the beginning of August, 1997. On blood test an increaseof the bilirubin level up to 34 mmol/L and GPT level up to 2.1mmol/hr.L. were found. During examination in the hospital from Aug. 15,1997, anti-HCV IgG and the replicative activity of the hepatitis C viruswere found.

[0543] Anamnesis vitae: The patient started using drugs at 14. To theexamination time he uses up to 2 g of heroin per day. He is at the stateof the narcotic abstinence.

[0544] Previous treatment was not conducted.

[0545] Immunomodulating therapy course with GSSG.Pt drugs: from Aug. 15,1997, to Sep. 7, 1997.

[0546] Patient's state after the treatment course completion: Thepatient's state is satisfactory. He noticed significant reduction ofweakness, no pains in the right under the ribs and in the joints. Aspatient said the narcotic abstinence state diminished almost withoutpain and in less time. Biochemical indices normalization and absence ofthe viral replicative activity was marked.

[0547] Conclusion: The immunomodulating therapy course with the GSSG.Ptdrug provided positive changes, which were indicated by: biochemical andserologic indices normalization; termination of HCV replication. Immuneindices and cytokine status parameters correlate to the infectiousprocess controlling and viral replication absence. Examination of thepatient's peripheral blood lymphocytes by liquid chromatography withmonoclonal antibodies to FasAg (CD95⁺) after the treatment revealed theCD95⁺ cell increase indicating activation of the programmed cell deathprocess in virus-infected cells. At supervision at one and three monthsafter the treatment stabilization of this state was noted.

[0548] Concomitant drug intoxication and an abstinence state atapplication of the GSSG.Pt drugs were corrected faster and were lessexcruciating for the patient.

[0549] The immunomodulating course with the GSSG.Pt drugs for chronichepatitis C with replicative activity and concomitant drug intoxicationhas provided the following results:

[0550] biochemical indices normalization in blood;

[0551] hematological indices normalization;

[0552] termination of HCV replication;

[0553] normalization of the immune blood indices and cytokine status;

[0554] apoptosis process induction in the peripheral blood lymphocytes;

[0555] rapid correction for the drug abstinence state;

[0556] stable therapeutic effect

EXAMPLE 14 Comparative Analysis of the GSSG and GSSG.Pt Effects onGrowth Development and Apoptotic DNA Degradation at Normal andTransformed Cells

[0557] The GSSG and GSSG.Pt effects on growth development and apoptoticDNA degradation at normal and transformed (HL-60) cells werecomparatively analyzed. To that end, GSSG and GSSG.Pt were incubated for24 hours with HL-60 myeloid line cells and normal human lymphocytesobtained from healthy volunteers' blood.

[0558] Venous blood of healthy volunteers was collected into heparinizedtest-tubes, which had been tested for endotoxin. A mononuclear fractionof blood leukocytes was obtained by centrifugation in ficoll-metrizoatgradient (Histopaque, Sigma). Cell concentration was adjusted to 2×10⁶cells per 1 ml of cell culture medium (RPMI 1640), containing 20 mMHEPES, 2 mM glutamine, 50 mg/mL gentamicin and 10% fetal calf serum.Cell viability was estimated by the Trypan blue exclusion method, thenthe cell suspension was placed into wells of 24- and 96-well microliterplates—250,000 cells per well.

[0559] The HL-60 myeloid line cells were grown in RPMI-1640 medium withaddition of 10% fetal calf serum. Cultivation was carried out in closedflasks, the medium volume was 12 mL, it was replaced every four days.Directly before testing the cell suspension in the fresh medium wasbrought into 24-well microliter plates (cell concentration for each wellwas 250.000 cells per well) and the tested articles—GSSG andGSSG.Pt—were added into the corresponding wells up to finalconcentration 100 mg/mL.

[0560] The effect testing for the testing articles was performed 24, 48hours after addition into the culture.

[0561] The analysis procedure involved the following: after 24-48 hourincubation there were calculated the total cell count and the dead cellcount by the Trypan blue exclusion method; afterwards the cellsuspension was centrifuged (at 12.000 g, in Eppendorf test-tubes during10 min). The cell pellet was frozen and kept at −70° C. before the DNAseparation. The DNA separation was conducted by Kirbi-Georgiyev phenolmethod. To the cell pellet there were added 0.5 ml of 10% SDS and 0.5 mlof TE-buffer containing 0.M EDTA and 0.01 M Tris-HCl with pH 8.0 (“A”buffer), and the pellet was resuspended by the tube shaking during 15min. Then the equal phenol amount was added adjusted by 0.01 M Tris-HClwith pH 8.0, the product was mixed during two min. and centrifuged inthe Eppendorf test-tubes at 12.000 g during 15 min. After thecentrifugation completion the upper water phase was phenol-treated onemore time. After the phenol treatment the upper water phase was twicetreated with phenol-chloroform mixture (1:1) that was mixed with thewater phase in equal amount. The water phase was separated bycentrifugation at 12.000 g during 10 min., pumped out and once mixedwith the equal chloroform amount. Afterwards it was centrifuged at12.000 g, the upper phase was separated, mixed with double amount ofdistilled ethyl alcohol frozen to −20° C. and kept for one night at −20°C. The DNA pellet was gathered by centrifugation at 12.000 g during 10min., the supernatant was removed and the pellet was washed by 200 mL of70% ethyl alcohol frozen to −20° C. during five min., centrifuged onemore time at the same conditions and afterwards the pellet was air-driedduring one hour. Then it was diluted in 10 mL of the A buffer; the DNAamount was determined by Dishe method and electrophoresis was performedin 2% agarose gel (agarose with NA grade made by “Pharmacia LKB,Biotechnology Inc” (Austria) was used). The DNA electrophoreticseparation was made in a block of 2% agarose gel in a device made by“Pharmacia LKB, Biotechnology Inc” (Austria). As a buffer solution 0.04M tris-HCl buffer, pH 7, containing 0.02 M of sodium acetate and 0.02 MEDTA was applied. Agarose (2%) was prepared at an electrode buffer. TheDNA samples (2-5 mg) were placed into the gel slots. The electrophoresiswas conducted with an electrical field intensity of 6 W/cm during threehours. The sample propulsion was observed due to bromine-phenol bluemotion. On the electrophoresis completion the gel block was taken out ofthe device and introduced into a tray with etidium bromide solution (3mg/mL H2O) for 30 minutes in dark place. After the incubation completionthe gel was rinsed with water and examined in the transmittedultraviolet radiation with the wave-length 254 nm at a transillunimiatormade by “Pharmacia LKB, Biotechnology Inc” (Austria). The gel wasphotographed by Zenit E camera with a red colour filter.

[0562] The study results are given in the Tables 28 and 29 and FIGS. 13and 14. As one can see in the Tables 28 and 29 data, the GSSG andGSSG.Pt influence on normal and transformed cells is of an alternativecharacter. GSSG and GSSG.Pt stimulated the normal cell proliferation(Table 28). The electrophoresis of DNA obtained from the normal cells(Table 28) revealed presence of only traceable quantities for apoptoticfragments at the background of homogenous high-molecular fractionhearacteristic for the viable cells.

[0563] In the contrary, in the myeloid origin (HL-60) cancer cellculture, the apoptosis activation and the cell division inhibition wereobserved due to the influence of both drugs(Table 29, FIG. 14). Thediversities of the effects were of the quantitative nature.

[0564] Thus, the dead HL-60 cell quantity after the GSSG impact wasreliably lower than the one after the GSSG.Pt effect (Table 29). Anevident indication for stronger impact efficacy (GSSG.Pt comparing toGSSG) on the HL-60 cells is the apoptotic fragment's character obtainedafter the electrophoretic analysis. The DNA electrophoresis of the HL-60cells non-incubated with the drugs demonstrated presence ofhighly-molecular, practically homogenous DNA characteristic for theviable cells (FIG. 14, band #2). The DNA electrophoresis of the cellsincubated during 24 and 48 hours with the GSSG.Pt and GSSG drugsrevealed the DNA oligonucleosomic degradation (FIG. 14, bands 1 and 3,respectively), i.e., an apoptotic ladder, that is indisputable sign forthe programmed cell death. However, the apoptotic ladder for the HL-60cells treated with GSSG contained considerable amount of the highmolecular DNA characteristic for the viable HL-60 cells (FIG. 14, band#1), whereas the high molecular DNA for the cells treated with GSSG.Ptwas virtually absent (FIG. 14, band #3).

[0565] The HL-60 cells are defective in the p53 gene (p53 gene deletion)and apoptosis induction through the GSSG and GSSG.Pt treatment can occuronly without involving the p53 product. Therefore, one can state thatthe GSSG and GSSG.Pt drugs activate an inner contour of the programmedcell death irrespectively of the p53 gene product. The p53 defect ispresent approximately at half of cancer pathology cases. Theexperimental results appeared to denote efficacy of GSSG and,especially, GSSG.Pt for these tumor chemotherapy.

[0566] Conclusion: The performed studies allowed to obtain dataindicating capacity of the GSSG and GSSG.Pt drugs to increase normalcell (lymphocytes) viability and, contrariwise, to induce apoptosis inransformed cells, i.e., exercise an antitumor activity. Besides, theGSSG.Pt activity regarding to the transformed cell apoptosis inductionexcelled significantly the one for the GSSG drug according to theobjective viability criterion, i.e., presence of the high molecular DNAthat appeared considerably after the GSSG treatment and was almostabsent after the GSSG.Pt treatment. Thus, the data obtained on cells ofthe myeloid line HL-60 defective in the p53 gene that is the crucialagent in apoptosis induction allows us to state that:

[0567] The GSSG and GSSG.Pt drugs induce the inner contour of theapoptosis development regardless of the p53 gene product;

[0568] The GSSG.Pt drug based on composite possesses higherchemotherapeutic activity (due to the apoptotic induction in thetransformed cells) versus the first generation drug GSSG.

EXAMPLE 15 Analysis of the GSSG.Pt Effects on Growth Development andApoptotic DNA Degradation at Normal and Transformed Cells Defective inthe p53 Antioncogene with Increased Ras-Gene Expression

[0569] The GSSG.Pt effects on growth development and apoptotic DNAdegradation of different lineage of transformed cells (HL-60, C-8, A-4)were comparatively analyzed depending on the p53 defect that is the keyfactor for the apoptosis development and the ras-gene that is amultipotent factor for cell reaction. The HL-60 cell culture is humancells of myeloluekosis origin defective in the p53 gene. Cell culturesC-8 and A-4 are transformed murine fibroblasts having a plasmid with theras gene and a gene of the E1a expression enhancement factor that is anadenovirus antigen fragment. At that, the A-4 cells are defective at thep53 gene and the C-8 ones contained the intact p53 gene. A donor bloodlymphocyte slip was used as a control of the human cells with the intactp53 gene.

[0570] The HL-60 (p53−−) myeloid line cells were grown in RPMI-1640medium with addition of 10% fetal calf serum. The cell suspensioncultivation was carried out in closed flasks, the medium volume was 12mL, it was changed every four days. Directly before testing the cellsuspension in the fresh medium was brought into 24-well microliterplates (cell concentration for each well was 250.000 cells per well) andthe tested article—GSSG.Pt—was added into the corresponding wells up tofinal concentrations 10-100 mg/mL.

[0571] Venous blood of healthy volunteers was collected into heparinizedtest-tubes, which had been tested for endotoxin. A blood leukocytemononuclear fraction was obtained by centrifugation in ficoll-metrizoatgradient (Histopaque, Sigma). Cell concentration was adjusted to 2×10⁶cells per 1 ml of cell culture medium (RPMI 1640), containing 20 mMHEPES, 2 mM glutamine, 50 mg/mL gentamicin and 10% fetal calf serum.Cell viability was estimated by the Trypan blue exclusion method, thenthe cell suspension was placed into wells of 24- and 96-well microliterplates—250,000 cells per well.

[0572] The murine transformed fibroblasts were grown in DMEM medium withaddition of 10% fetal calf serum. The cell suspension cultivation wascarried out in closed flasks, the medium volume was 12 mL, it waschanged every four days. Directly before testing the cell suspension inthe fresh medium was brought into 24-well microliter plates (cellconcentration for each well was 50.000 cells per well) and the testedarticle—GSSG.Pt—was added into the corresponding wells up to finalconcentrations 10-100 mg/mL.

[0573] The effects testing for the testing articles was performed 24, 48hours after introduction into the culture.

[0574] The analysis procedure involved the following: after 24-48 hourincubation, the total cell count and the dead cell count were calculatedby the Trypan blue exclusion method; afterwards the cell suspension wascentrifuged (at 3.000 g, in Eppendorf test-tubes during 10 min). Thecell pellet was frozen and kept at −70° C. before the DNA separation.The DNA separation was conducted by Kirbi-Georgiyev phenol method. Tothe cell pellet there were added 0.5 ml of 10% SDS and 0.5 ml ofTE-buffer containing 0.1M EDTA and 0.01 M Tris-HCl with pH 8.0 (“A”buffer), and the pellet was resuspended by the tube shaking during 15min. Then the equal phenol amount was added adjusted by 0.01 M Tris-HClwith pH 8.0, the product was mixed during two min. and centrifuged inthe Eppendorf's test-tubes at 12.000 g during 15 min. After thecentrifugation completion the upper water phase was phenol-treated onemore time. After the phenol treatment the upper water phase was twicetreated with phenol-chloroform mixture (1:1) that was mixed with thewater phase in equal amount. The water phase was separated bycentrifugation at 12.000 g during 10 min., pumped out and once mixedwith the equal chloroform amount. Afterwards it was centrifuged at12.000 g, the upper phase was separated, mixed with double amount ofdistilled ethyl alcohol frozen to −20° C. and kept for one night at −20°C. The DNA pellet was gathered by centrifugation at 12.000 g during 10min., the supernatant was removed and the pellet was washed by 200 mL of70% ethyl alcohol frozen to −20° C. during five min., centrifuged onemore time at the same conditions and afterwards the sediment wasair-dried during one hour. Then it was diluted in 10 mL of the A buffer;the DNA amount was determined by Dishe method and electrophoresis wasperformed in 2% agarose gel (agarose with NA grade made by “PharmaciaLKB, Biotechnology Inc” (Austria) was used). The DNA electrophoreticseparation was made in a block of 2% agarose gel in a device made by“Pharmacia LKB, Biotechnology Inc” (Austria). As a buffer solution 0.04M tris-HCl buffer, pH 7, containing 0.02 M of sodium acetate and 0.02 MEDTA was applied. Agarose (2%) was prepared at an electrode buffer. TheDNA samples (2-5 mg) were placed into the gel slots. The electrophoresiswas conducted with an electrical field intensity of 6 W/cm during threehours. The sample propulsion was observed due to bromine-phenol blueindicator motion. On the electrophoresis completion the gel block wastaken out of the device and introduced into a tray with etidium bromidesolution (3 mg/mL H₂O) for 30 minutes in dark place. After theincubation completion the gel was rinsed with water and examined in thetransmitted ultraviolet radiation with the wave-length 254 nm at atransilluminator made by “Pharmacia LKB, Biotechnology Inc” (Austria).The gel was photographed by Zenit E camera with a red color filter.

[0575] The study results are given in the Tables 30, 31, 32, 33. As isevident from the Table 31, GSSG.Pt induces apoptosis in the HL-60 cellculture defective at the p53 gene. The effect was rather developed atthe concentration of 10, however, it was more evident at 100 mg/mL. Alsothere were observed conglomeration of DNA apoptotic fragments multipleof DNA nucleosome size that is the indisputable sign of programmed celldeath.

[0576] In contrary, GSSG.Pt made stimulating effect on normal cells,i.e., there was certain proliferation observed (Table 30).Electrophoresis of DNA obtained from normal cells exhibited that it wasrepresented by a homogenous high-molecular fraction characteristic forviable cells.

[0577] Thus, differences of the drug action on normal and transformedcells were basically divergent. Mechanism for the GSSG.Pt divergentaction might be conditioned by activation of the p53-independentapoptotic pathway through the ras-signal-transducing system. Theras-system is capable to stimulate cell proliferation anddifferentiation through the mitogenic factor cascade and induceapoptosis (programmed cell death) through another cell signal cascade.

[0578] To check it there were compared the GSSG.Pt effects on murinefibroblasts with enhanced ras-gene expression but different in theintact p53 gene (wild type) presence—cells C-8(p53++), or the p53 geneabsence (a genetic defect)—(cells A-4(p53−−)). As one can see from theTables 32 and 33, the GSSG.Pt effect appeared in both cell lineages. Theapoptosis induction was significantly exhibited. It indicates activationof the p53-independent apoptotic pathway. Presence of the activated rasgene in both of the cell lineages can be an explanation for theapoptotic exhibition that was even more marked in the transformedfibroblasts than in the HL-60 cells. It confirmed the apoptosisinduction through the ras-signal-transducing system. The GSSG.Pt effecton the A-4 and C-8 cells was not different at concentration of 10 mg/mL.The significantly superior GSSG.Pt effect on the A-4 cells comparing tothe C-8 cells was noted at the concentration 100 mg/mL. The p53-proteinabsence appeared even to enhance the ras-signal-transducing pathway forapoptosis induction in tumor cells.

[0579] The p53 gene defect occurs in approximately half of the cancerdisease cases. The results of these experiments can imply the GSSG.Pteffectiveness for chemotherapy of these tumors.

[0580] Conclusion: The data from these studies indicate the capacity ofthe GSSG.Pt drug to increase normal cell (lymphocytes) viability and,contrariwise, to induce apoptosis in transformed cells, i.e., exercisean antitumor activity. Besides, the GSSG.Pt activity regarding to thetransformed cell (defective at the p53 gene) apoptosis induction evenexcelled the one for the cells with the intact p53 gene at the high drugconcentration. According to the objective viability criterion, i.e.,presence of the high molecular DNA that appeared in considerable amountsafter the GSSG.Pt treatment of the normal donor cells (take from thelymphocyte slip), the cell death was practically absent whereas in caseof the GSSG.Pt impact on transformed cells there was observed the DNAapoptotic degradation, i.e., the sign of the irreversible apoptoticdeath induction even in the cells defective at the p53 gene (with thestimulation especially). Previously, in the Example 9 there was shownactivation of the cytokine range after the GSSG.Pt treatment.Considering that the cytokine action may be determined with theras-signal-transducing pathway activation the cytokine stimulation canalso cause the antitumor effect through an interaction with theras-protein.

EXAMPLE 16 Analysis of the GSSG.Pt Effects on Growth Development andApoptotic DNA Degradation at Murine Transformed Cells Cultures Defectiveat Antioncogenes

[0581] The GSSG.Pt effects on growth development and apoptotic DNAdegradation of transformed fibroblasts of a cell lineage with activatedras-gene but an intact p21 gene (p21++—C-8 cells) and murine celllineage with knockout p21 gene (p21−−).

[0582] The p21++ lineage cells are murine transformed fibroblasts havinga plasmid with the ras gene and a gene of the E1a expression enhancementfactor that is an adenovirus antigens fragment but have intact p53 andp21 genes. The p21 (−−) lineage has the intact p53 and ras genes but itis defective at the p21 gene. It allows evaluating the GSSG.Pt impact onthe apoptosis induction in conditions of impaired regulation for thecell division G1 phase. The cell cultures were grown in DMEM medium withaddition of 10% fetal calf serum. The cell suspension cultivation wascarried out in closed flasks, the medium volume was 12 mL, it waschanged every four days. Directly before testing, the cell suspension inthe fresh medium was brought into 24-well microliter plates (cellconcentration for each well was 50.000 cells per well) and the testedarticle—GSSG.Pt—was added into the corresponding wells up to finalconcentrations 100 mg/mL.

[0583] The effect testing for the testing articles was performed 24, 48hours after introduction into the culture.

[0584] The analysis procedure involved the following: after 24-48 hourincubation, the total cell count and the dead cell count were calculatedby the Trypan blue exclusion method; afterwards the cell suspension wascentrifuged (at 3.000 g, in Eppendorf test-tubes during 10 min). Thecell pellet was frozen and kept at −70° C. before the DNA separation.The DNA separation was conducted by Kirbi-Georgiyev phenol method. Thecell lysis was performed by addition of 0.5 ml of 10% SDS. The equalphenol amount was added there adjusted by 0.01 M Tris-HCl with pH 8.0,the product was mixed during two min. and centrifuged in the Eppendorftest-tubes at 13.000 g during 15 min. The phenol deproteinization wasrepeated twice. Afterwards the water phase was twice treated withphenol-chloroform mixture (1:1) and once with chloroform. Nucleic acidswere precipitated by the addition of two volumes of 96% ethyl alcohol at20° C. overnight. The DNA was gathered by centrifugation at 13.000 gduring 30 min., decanted, washed with 70% ethyl alcohol and air-dried.After the drying the precipitate was dissolved in TE buffer. The DNAconcentration was determined in the obtained solution (by Dishe method).Fractional content of the nucleic acids was determined byelectrophoresis in 2% agarose gel (agarose with NA grade made by“Pharmacia LKB, Biotechnology Inc” (Austria) was used). The DNAelectrophoretic separation was made in a block of 2% agarose gel in adevice made by “Pharmacia LKB, Biotechnology Inc” (Austria). Theelectrophoresis was performed in TAE buffer pH 7.4 (0.04 M tris, 0.02 Mof sodium acetate, 0.02 M of EDTA) at an electrical field intensity of 6W/cm during three hours. The sample propulsion was observed due tobromine-phenol blue indicator motion. The electrophoresis results wereexamined in the transmitted ultraviolet radiation (γ=254 nm) at atransilluminator made by “Pharmacia LKB, Biotechnology Inc” (Austria)after the gel dying with etidium bromide solution (5 mg/ml).

[0585] The study results are given in the Tables 34, 35. As it isevident from the Table 34, GSSG.Pt induces apoptosis in the p21++ cellculture with activated ras-gene manifested by conglomeration of DNAapoptotic fragments multiple of DNA nucleosome size.

[0586] In the p21 (−−) cells having non-active ras-signal-transducingsystem but defective at the control of the cell cycle delay in the GIphase GSSG.Pt induced apoptosis to less extent that in p21++ cells(Tables 34, 35). It might be implicated by the fact that the ras-systemis capable of stimulatin cell proliferation and differentiation throughthe mitogenic factors' cascade and inducing apoptosis (programmed celldeath) through another cell signals' cascade. Absence of the p21 geneexpression can cause changes in interrelations of specific cascades ofthe ras-signal-transducing pathway and, thereupon, lessen the apoptoticstimulation by the drug in cases of p21 gene defects.

[0587] The p21 gene defect does not often occur in oncological diseasesthat considering the performed experiments' issues can imply the GSSG.Pteffectiveness for chemotherapy of these tumors except cases with lowereffectiveness at tumors with mutations in p21 gene.

[0588] Conclusion: The performed studies allowed obtaining dataindicating capacity of the GSSG.Pt drug to induce apoptosis actively intransformed cells, i.e., exercise an antitumor activity. The enhancedras-gene expression facilitates the apoptosis induction in thetransformed cells C-8 indicating implementation of the GSSG.Pt antitumoractivity through the ras-signal-transducing system. The lesser drugeffect in case of p21 gene expression absence can be determined by theredistribution of factors in mitogenic and apoptotic cascades of theras-signal-transducing pathway.

[0589] Nevertheless, presence of the DNA apoptotic degradation, i.e.,the sign of the irreversiblc apoptotic death induction, is found afterthe GSSG.Pt action even in p21-defective cells.

[0590] The aforesaid allows us to state that the GSSG.Pt composite-baseddrug:

[0591] Realizes chemotherapeutic activity regarding thetumor-transformed cells through induction of the ras-dependent apoptoticpathway;

[0592] Induces the apoptosis' development in the Lumor-transformed cellsincluding p21-defective cells.

EXAMPLE 17 Therapeutical Effect of the GSSG.Pt Vanadium Salt in Patientwith Diabetes Mellitus

[0593] No.7

[0594] Gender: female

[0595] Age: 37

[0596] Out-patient card: No. 63

[0597] Diagnosis: Diabetes mellitus. Insulin-independent type—type II.Diabetic angiopathy, grade IV.

[0598] Anamnesis morbi: Firstly the high blood sugar was revealed in theage of 31. In October 1994, the patient was admitted into theEndocrinologic Department of the Hospital #16. There were diabetes casesin the patient's hereditary history on the mother's side. The diseasedeveloped gradually, the blood glucose level fluctuated from 12.1 to15.7 mmol/L, glucosuria to 7%.

[0599] Previous treatment: At the onset of the disease, the patient wasfrequently admitted to hospitals; in 1994 during six months she wasadministered with Insulin-lente S.N.C.-32 IU and peroral hypoglycemicagents of the sulfonylureas group; then Insulin was cancelled and thepatient used antidiabetic agents—Bucarban, Diabeton. The blood glucosechanged from 10.7 to 15.4 mmol/L, glucosuria 3-7%.

[0600] In January 1998 the patient was taken to the hospital and thefollowing treatment regime was administered: Diabeton 0.08 g —2 tabtwice a day, Glucobay 0.1×3 times per day.

[0601] The blood glucose content: 9 12.6 mmol/L 12 12.0 mmol/L 14 15.3mmol/L 17 19.1 mmol/L 6 11.5 mmol/L

[0602] Glucosuria—to 3%

[0603] Because of the previous treatment inefficacy, it was decided touse the drug of the GSSG.Pt vanadium salt.

[0604] Hospital treatment regime for the GSSG.Pt vanadium salt: fromJan. 17, 1998.

[0605] During 10 days once a day—intravenous injections of the GSSG.Ptvanadium salt (daily dose—0.01-0.5 mg/kg).

[0606] During the following 10 days—intravenous injections of theGSSG.Pt vanadium salt every other day (daily dose—0. 01-0. 5 mg/kg).

[0607] During the following 10 days (the third decade)—intramuscularinjections of the GSSG.Pt vanadium salt once a day (daily dose—0.01-0.5mg/kg).

[0608] The GSSG.Pt vanadium salt treatment was performed along withchanged treatment regime by Diabeton and Glucobay. Diabeton wasadministered 0.08 twice a day, Glucobay 0.05×3 times a day.

[0609] The blood glucose restored to normal values and did not exceed8.2-10.6 mmol/L after meals. After the discharge the patient receivedthe GSSG.Pt vanadium salt treatment as an out-patient during one month.

[0610] Ambulant treatment regime for the GSSG.Pt vanadium salt:

[0611] For one month there were administered intramuscular injections ofthe GSSG.Pt vanadium salt once a day (daily dose—0.01-0.5 mg/kg).

[0612] During two sequential months of treatment with the GSSG.Ptvanadium salt drug there were noted two episodes of the glucose contentdecrease to 1.5-2.5 mmol/L, thereat the antidiabetic drugs dosages werelowered. After two months of the treatment, Glucobay was cancelled.

[0613] Patient's state after the treatment course completion: After fourmonths of the treatment with administration of the GSSG.Pt vanadium saltdrug as a support for the basic therapy, the patient's general stateimproved significantly and was evaluated as satisfactory. Development ofhematologic, biochemical, immunologic blood indices is provided in theTables 36, 37.

[0614] Comments to the indices development of glucose, cAMP/cGMP,thiol-disulfide ratio and other analyzed parameters.

[0615] The glucose content development is an integrative criterion forthe impact effectiveness of the antidiabetic drugs. The combined therapyby the peroral antidiabetic drugs (Diabeton and Glucobay) along with theGSSG.Pt vanadium salt drug called forth the blood glucose normalization,decrease of the Diabeton therapeutic dosage and the Glucobaycancellation. Character of the hematologic, biochemical and immuneparameter changes indicated restoration of the metabolism and thesystemic reactions in general that does not allow to determine theantidiabetic constituent for the GSSG.Pt vanadium salt drug. Thefollowing indices were used—cAMP/cGMP and thiol-disulfide ratio(TDR)—that allowed to characterize basically a molecular mechanism ofthe GSSG.Pt vanadium salt drug action on cellular reaction complexstabilizing the blood glucose level. In particular, the keyintracellular messengers—cAMP and cGMP—determine intensity of theglucose flow into intracellular metabolic processes. At that,cGMP-dependent enzymatic cellular systems determine the cellular glucoseintake intensity. In its turn, the cGMP level is determined by thecellular oxidative potential. In particular, oxidants increase the cGMPlevel, in the contrary, antioxidants perform depressive action on itscontent. Thus, the intracellular thiol-disulfide ratio (TDR) reflectingthe balance of the anti- and pro-oxidants determines value of theintracellular cAMP/cGMP index. Taking into account an organism as awhole we can analyze these indices in blood as integrative internalmedium interrelated with the glucose level fluctuations, because theglucose metabolism is an indefeasible constituent for functioning of allcell types at our organism along with thiol-disulfide metabolism andcyclic nucleotide systems.

[0616] The results obtained indicated the glucose-lowering influence ofthe GSSG.Pt vanadium salt drug. Besides, this effect follows the cGMPcontent increase development (lowering of the cAMP/cGMP index) and theTDR value decrease (increase of the thiol oxidized forms). Consideringregulatory possibilities of the thiol oxidized forms and the cGMP inblood glucose regulation, one can state the basic mechanism of theregulatory hypoglycemic effect of the GSSG.Pt vanadium salt drug. Interalia, the regulatory impact of the GSSG.Pt vanadium salt drug on thecellular redox contour, provides an increase of the oxidant constituentlevel that, in its turn, redistributes balance in the cyclic nucleotidesystem in favor of cGMP (guanylat-cyclase induction and inhibition ofphosphodiesterase, cGMP synthesis and destruction enzymes,respectively). The cGMP regulatory impact stimulates the glucosetransport processes into insulin-dependent tissues calling forth theblood glucose decrease.

[0617] Conclusion: Application of the GSSG.Pt vanadium salt drug in thescheme for the combined diabetes treatment allowed to obtain thefollowing therapeutic effects:

[0618] the quality of life improvement and the blood glucose levelstabilization;

[0619] dosage decrease for the administered glucose-lowering drugs;

[0620] restoration to normal values of the hematologic, biochemical andimmunologic indices.

[0621] Those skilled in the art would readily appreciate that allparameters listed herein are meant to be examples and that actualparameters will depend upon the specific application for which themethods and apparatus of the present invention are used. It is,therefore, to be understood that the foregoing embodiments are presentedby way of example only and that, within the scope of the appended claimsand equivalents thereto, the invention may be practiced otherwise thanas specifically described. TABLE 1 Variation of the GSSG content (μg/mL)in the blood serum of CBA mice after the GSSG drug intravenousintroduction in different doses, M ± m GSSG dose Time intervals, min 2mg/kg 20 mg/kg GSSG content, μg/mL 0 0.3 ± 0.02 0.42 ± 0.03  1 12.4 ±1.3*   130 ± 12.2*  2 9.4 ± 0.87 89.5 ± 9.1*  5  3.3 ± 0.28* 35.6 ±4.2*  10  0.6 ± 0.05* 4.2 ± 0.37* 20 0.44 ± 0.4  3.9 ± 0.31* 40 0.32 ±0.05* 3.4 ± 0.32* 60 0.3 ± 0.05 3.3 ± 0.27*

[0622] TABLE 2 Variation of thc GSSG content (μg/mL) in the blood serumof CBA mice after the GSSG•Pt drug intravenous introduction in differentdoses M ± m. GSSG•Pt dose Time intervals, min 2 mg/kg 20 mg/kg GSSGcontent, μg/mL 0  0.2 ± 0.01 0.37 ± 0.04   1 56.4 ± 5.3  572 ± 56.3* 2 33.2 ± 0.24* 345 ± 32.1* 5 23.3 ± 1.4  227 ± 20.3* 10 13.2 ± 1.5* 129 ±13.5* 20 10.6 ± 1.2* 107 ± 10.2* 40  9.4 ± 0.91* 98 ± 9.3* 60  9.1 ±0.93 94 ± 9.3*

[0623] TABLE 3 The GSSG content in the blood serum and different organsof the CBA mice after the GSSG drug intravenous introduction in dose of2 μg/mL, M ± m. Time interval, Blood serum, Liver Kidneys SpleenLymphocytes, min. μg/mL μg/gram of tissue f × 10⁵ of cells 0 0.3 ± 0.029.1 ± 0.9 5.3 ± 0.6 3.3 ± 0.4  8.9 ± 0.96 1 12.4 ± 1.3*  402 ± 41*   212± 22.3*   137 ± 0.14*  356 ± 32.1 2 9.4 ± 0.87   292 ± 33.2*  105 ± 11.498.6 ± 9.5    157 ± 16.2* 5  3.3 ± 0.28* 94.3 ± 9.7  74.3 ± 7.6* 67.5 ±6.4* 89.3 ± 8.4* 10  0.6 ± 0.05* 17.2 ± 1.4* 16.3 ± 1.4  12.8 ± 3.5*21.8 ± 2.6  20 0.44 ± 0.4   9.5 ± 0.9*  6.1 ± 0.6*  4.1 ± 0.56 10.3 ±1.1* 40 0.32 ± 0.05* 9.3 ± 0.9  6.7 ± 0.5*  3.7 ± 0.32 9.6 ± 0.9 60 0.3± 0.05  8.9 ± 0.9*  5.5 ± 0.47*  3.5 ± 0.4*  8.7 ± 0.8*

[0624] TABLE 4 The GSSG content in the blood serum and different organsof the CBA mice after the GSSG•Pt drug intravenous introduction in doseof 2 μg/mL, M ± m. Time Blood interval, serum, Liver Kidneys SpleenLymphocytes min. μg/mL μg/gram of tissue f × 10⁵ of cells 0  0.2 ± 0.019.1 ± 0.9   5.3 ± 0.6   3.3 ± 0.4   8.9 ± 0.96  1 56.4 ± 5.3  2613 ±220.3* 1842 ± 194.1* 936 ± 92.7* 2247 ± 229.3  2  33.2 ± 0.24* 1315 ±132.4* 832 ± 87.7* 447 ± 42.7  1216 ± 123.3* 5 23.3 ± 1.4  754 ± 76.8 449 ± 45.6* 219 ± 23.6* 715 ± 82.4* 10 13.2 ± 1.5* 382 ± 33.7* 227 ±23.2* 107 ± 10.3* 306 ± 33.3* 20 10.6 ± 1.2* 378 ± 31.2* 216 ± 22.3  105± 10.1* 296 ± 28.2* 40  9.4 ± 0.91* 369 ± 29.3* 213 ± 21*   103 ± 10.1*289 ± 24.1  60  9.1 ± 0.93 367 ± 32.9* 215 ± 21.2* 102 ± 10.4  287 ±20.8*

[0625] TABLE 5 Activity of the enzymes participating in the thiolmetabolism at the intact CBA mice, M ± m. Enzyme activity Liver KidneysSpleen Thymus 1. glutathione- 65 ± 3 89 ± 5 31 ± 4 11 ± 3 reductase⁽¹⁾2. glutathione- 32 ± 4 29 ± 6 12 ± 2  8 ± 2 peroxidase⁽²⁾ 3.glutathione-S- 242 ± 14 310 ± 18 87 ± 6 41 ± 4 transferase⁽²⁾ 4.γ-glutamyl- 46 ± 8 96 ± 6 12 ± 3 14 ± 2 transpeptidase⁽³⁾

[0626] TABLE 6 Activity changes for the enzymes participating in thethiol metabolism at the CBA mice after the GSSG drug intravenousintroduction in dose of 20 μg/mL, M ± m. Enzyme activity Liver KidneysSpleen Thymus 1. glutathione- 128 ± 13* 169 ± 12* 74 ± 6* 24 ± 4*reductase⁽¹⁾ 2. glutathione- 54 ± 6* 46 ± 8* 18 ± 3* 14 ± 2*peroxidase⁽²⁾ 3. glutathione-S- 442 ± 18* 510 ± 14* 162 ± 8*  81 ± 6*transferase⁽²⁾ 4. γ-glutamyl- 96 ± 7* 189 ± 20* 23 ± 4* 28 ± 3*transpeptidase⁽³⁾

[0627] TABLE 7 Activity changes for the enzymes participating in thethiol metabolism at thc CBA mice after the GSSG•Pt drug intravenousintroduction in dose of 20 μg/mL, M ± m. Enzyme activity Liver KidneysSpleen Thymus 1. glutathione- 67 ± 3* 91 ± 4 37 ± 4 13 ± 2* reductase⁽¹⁾2. glutathione- 33 ± 4   31 ± 4* 14 ± 2 9 ± 1 peroxidase⁽²⁾ 3.glutathione-S- 251 ± 16  324 ± 22 93 ± 7 44 ± 5  transferase⁽²⁾ 4.γ-glutamyl- 49 ± 5* 99 ± 8  15 ± 2* 16 ± 1  transpeptidase⁽³⁾

[0628] TABLE 8 GSSGPt effect on in vitro cytokine production by humanmononuclear leukocytes. (M ± m) Cytokine GSSGPt (μg/mL) productionControl (pg/mL) 0.5 5.0 50 500 5000 (RPMI) IL-1β 56.0 ± 9.1  88.5 ±13.5*  202 ± 24.9*  275 ± 39.3*  259 ± 36.8  46.0 ± 6.8  IL-2 87.2 ±7.5   123 ± 10.6*  234 ± 21.5*  310 ± 32.1*  348 ± 29.4*   51 ± 5.4 IL-6  430 ± 55.6  550 ± 61.3* 1810 ± 205*  2103 ± 132*  2518 ± 264*  129 ± 12.4 INF-α  130 ± 14.9  109 ± 12.1*  407 ± 51.4*  514 ± 56.2* 811 ± 64.1  98.3 ± 14.0 TNF-α 99.1 ± 11.6   314 ± 44.7   813 ± 90.8*1525 ± 163*  1900 ± 206*  88.7 ±  9.3

[0629] TABLE 9 GSSGPt effect on in vitro cytokine production by humanmononuclear leukocytes. (M ± m) Cytokine GSSGPt μg/mL) productionControl (pg/mL) 0.5 5.0 50 500 5000 (RPMI) IL-1β 83.0 ± 7.8   212 ±31.7*  511 ± 55.1*  650 ± 67.1*   620 ± 61.3*  46.0 ± 6.8  IL-2  117 ±11.5  263 ± 20.6*  532 ± 53.5*  703 ± 72.0*   848 ± 89.4    51 ± 5.4 IL-3 436 ± 43   543 ± 55.2*  754 ± 74.5*  965 ± 87.4*  1024 ± 108*   206 ± 22.4 IL-4  160 ± 14.9  143 ± 14.1*  124 ± 13.3*  107 ± 10.1*  84.9 ± 7.1   175.3 ± 16.0 IL-6  851 ± 111  1680 ± 207*  3859 ± 425* 4007 ± 419*   4035 ± 518*    129 ± 12.4 IL-8  123 ± 13.7  134 ± 13.2* 138 ± 13.5*  140 ± 13.6    143 ± 13.9*   114 ± 11.3 IL-10  174 ± 16.4 153 ± 14.9*  126 ± 11.4*  109 ± 9.7*    94 ± 9.2    206 ± 19.2 IL-12 146 ± 13.2  164 ± 15.6*  186 ± 17.9*  208 ± 19.5    227 ± 21.4*   115 ±10.3 INF-α  150 ± 14.9  176 ± 17.1*  468 ± 69.3*  905 ± 141     849 ±1121   98.3 ± 14.0 IFN-γ  156 ± 14.8  175 ± 16.9*  194 ± 18.5*  205 ±21.4*   267 ± 25.1*   132 ± 11.4 TNF-α  318 ± 47.8  502 ± 86.4* 1308 ±164*  2100 ± 294*   2640 ± 355    88.7 ± 9.3  TNF-γ  167 ± 15.8  386 ±37.5*  584 ± 57.7*  796 ± 78.4*  1063 ± 10*     109 ± 9.5 

[0630] TABLE 10 Effect of the GSSG and GSSGPt on cytokines' andhemopoietic factors' production by splenocytes, bone marrow and bloodcellular indices, and immune response to SRBC in cyclophosphamidetreated mice. (M ± m) Intact animals Cyclophosphamide-treated animalsParameter N Normal saline Normal saline GSSG GSSGPt Blood leukocytecount, 10 11.9 ± 1.81   4.7 ± 1.25*  8.5 ± 0.81◯  12.4 ± 1.2◯  10⁹/LBlood lymphocyte count, 10  7.4 ± 0.85   3.1 ± 0.56*  6.0 ± 1.28◯  6.9 ±1.04◯ 10⁹/L Bone marrow nucleated 10 53.7 ± 8.7   23.8 ± 5.0*   45.4 ±3.9 ◯  >62.3 ± 4.7◯  cell number, 10⁶/L SRBC agglutinin titer 10 5.33 ±0.74   1.47 ± 0.35*  3.08 ± 0.59◯  5.42 ± 0.54◯ (log₂) IL-1β 10   54 ±3.7   10.2 ± 1.6*   23.4 ± 2.5◯   49.7 ± 5.2◯  IL-2 10 76 ± 6.8 14.6 ±1.3*   35 ± 4.0◯  74.1 ± 7.2◯ IL-3 10  178 ± 16.5   34.5 ± 3.7*   72.4 ±6.9◯    174 ± 16.8◯ IL-4 10   89 ± 18.9  113.8 ± 4.6*  105.8 ± 7.6◯   87 ± 17.4◯ IL-6 10   98 ± 8.7   19.6 ± 1.8*   42.7 ± 4.1◯   95.7 ±9.4◯  IL-8 10  102 ± 9.5   23.4 ± 2.4*   52.6 ± 4.9◯   97.8 ± 8.9◯ IL-10 10 86.8 ± 17.3  137.8 ± 14.9* 126.9 ± 15.4◯  84.0 ± 8.9◯  IL-12 10  96 ± 8.5   18.7 ± 1.7*   38.3 ± 3.6◯   92.5 ± 8.5◯  INF-α 10  113 ±11.2   26.4 ± 2.5*   57.2 ± 5.2◯  109.5 ± 9.7◯  IFN-γ 10  126 ± 11.9  24.8 ± 2.2*   49.6 ± 4.1◯  118.6 ± 12.0◯ TNF-α 10   93 ± 8.5   17.4 ±1.6*   36.1 ± 3.2◯   89.7 ± 9.1◯  TNF-γ 10  115 ± 10.6   22.7 ± 2.5*  47.4 ± 5.1◯  111.3 ± 12.3◯ GM-CSF 10  180 ± 14.2   48.2 ± 7.2*  119.5 ±13.4◯ 178.2 ± 18.1◯ G-CSF 10  150 ± 13.7   26.7 ± 3.1*    67 ± 6.1◯ 148.4 ± 14.2◯ M-CSF 10  130 ± 10.03  34.2 ± 2.7*    71 ± 6.9◯  125.7 ±11.4◯

[0631] TABLE 11 General blood analysis of the male rats with cytopeniaafter the precursory cyclophosphamide introduction in the dose 50 mg/kgand the treatment with GSSG•Pt in the dose 5 mg/kg, ±mCyclophosphamide-treated Intact animals animals Parameter Normal salineNormal saline GSSG•Pt Day 10 Hemoglobin, g/dL 13.3 ± 0.4 10.3 ± 0.3*12.7 ± 0.3* Hematocrit, g % 48.4 ± 0.9 36.2 ± 0.8* 45.3 ± 1.1*Erythrocytes, 10¹²/L  6.1 ± 0.2  35 ± 0.3*  5.5 ± 0.3* Color index ( )pg17.8 ± 0.3 15.5 ± 0.3* 16.5 ± 0.5* ESR, mm/hr.  5.3 ± 0.2  6.6 ± 0.3* 5.5 ± 0.2* Platelets, 10⁹/L 808 ± 36 342 ± 52* 735 ± 42* Leukocytes,10⁹/L  7.8 ± 0.1  1.2 ± 0.1*  5.6 ± 0.2* Stab neutrophils, % 0 0 0Segmented neutrophils, 14.4 ± 1.0 58.2 ± 4.6* 19.2 ± 2.5* % Basophils, %0 0 0 Lymphocytes, % 82.0 ± 1.5 34.7 ± 0.9* 50.0 ± 0.8* Eosinophils, % 1.3 ± 0.3 0  2.0 ± 0.3* Monocytes, %  2.1 ± 0.2  7.9 ± 0.7*  2.8 ± 0.3*Plasma cells, % 0 0  2.0 ± 0.2* Reticulocytes, % 2 1 2 Normocytes, % 0 01 Polychromatic cells, % 0 2 1 Day 15 Hemoglobin, g/dL 14.3 ± 0.3 10.0 ±0.3* 13.0 ± 0.3* Hematocrit, g % 49.7 ± 1.0 30.4 ± 1.0* 46.7 ± 0.7*Erythrocytes, 10¹²/L  6.2 ± 0.1  3.4 ± 1.0*  5.7 ± 0.2* Color index ( )pg 18.2 ± 0.2 14.7 ± 0.2* 17.0 ± 0.3* ESR, mm/hr.  5.3 ± 0.2  7.6 ± 0.2* 5.6 ± 0.3* Platelets, 10⁹/L 824 ± 41 225 ± 49* 773 ± 40* Leukocytes,10⁹/L  8.0 ± 0.1  1.2 ± 0.21*  5.5 ± 0.3* Stab neutrophils, % 0 0 0Segmented neutrophils, 13.7 ± 1.7 42.2 ± 2.5* 18.0 ± 2.2* % Basophils, %0 0 0 Lymphocytes, % 77.3 ± 2.0 35.2 ± 3.5* 71.0 ± 1.8* Eosinophils, % 1.0 ± 0.3 0 0 Monocytes, %  2.1 ± 0.2  4.5 ± 0.5*  2.2 ± 0.3* Plasmacells, % 0 0  2.0 ± 0.2* Reticulocytes, % 2 1 1 Normocytes, % 0 0 0Polychromatic cells, % 0 3 1

[0632] TABLE 12 Myclogram of the male rats with cytopenia after theprecursory cyclophosphamide introduction in the dose 50 mg/kg and thetreatment with GSSG•Pt in the dose 5 mg/kg, ±m Intact animalsCyclophosphamide-treated animals Parameter Normal saline Normal salineGSSG•Pt Day 10 Cells' quantity, 71.4 ± 5.5  33.4 ± 4.2*  63.0 ± 7.4* 10¹¹/L Non-differentiated 0.4 ± 0.1  0* 1.0 ± 0.2* blasts, %Proerythroblasts, % 0 0 0 Erythroblasts, % 0.8 ± 0.2 0.3 ± 0.1* 1.8 ±0.4* Pronormoblasts, % 0.6 ± 0.1 0.2 ± 0.1* 0.5 ± 0.1* Basophilic 7.5 ±1.0 3.2 ± 0.4* 6.2 ± 1.08 normoblasts, % Polychromatic 8.4 ± 1.2 3.1 ±0.8* 16.5 ± 2.2*  normoblasts, % Eosinophilic 5.8 ± 1.2 2.3 ± 0.6* 8.3 ±2.0* normoblasts, % Red blood 0.56 ± 0.09 0.18 ± 0.08* 0.75 ± 0.22 mitosis count, % Myeloblasts, % 2.1 ± 0.5 1.2 ± 0.02 3.6 ± 0.6*Promyelocytes, % 4.0 ± 0.5 1.8 ± 0.5  5.6 ± 0.8* Myclocytes, % 6.1 ± 0.73.4 ± 1.1* 7.5 ± 0.88 Metamyelocytes, % 8.5 ± 1.2 4.5 ± 0.8* 6.7 ± 1.1*Stab neutrophils, % 11.6 ± 1.5  4.2 ± 1.0* 9.9 ± 1.4  Segmented 16.7 ±3.5  37.4 ± 5.2*  44.3 ± 5.0*  neutrophils, % Eosinophils, % 7.2 ± 1.23.1 ± 1.0* 7.0 ± 1.2* Basophils, % 0 0  0.1 ± 0.05* White blood 0.44 ±0.10 0.15 ± 0.04* 0.58 ± 0.14  mitosis count, % Prolymphocytes, % 0 00.5 ± 0.2* Lymphocytes, % 8.1 ± 0.7 2.2 ± 0.5* 16.4 ± 2.1*8 Plasmacells, % 1.3 ± 0.3 0 2.4 ± 0.2* Promonocytes, % 0 0 0.1 Monocytes, % 0.8± 0.2  0.1 ± 0.05* 0.5 ± 0.2* Reticulocytes, % 2.6 ± 0.7 0.9 ± 0.3  3.5± 1.2* Megakaryocytes and 0.45 ± 0.08 0.12 ± 0.08* 0.62 ± 0.11*megakaryoblasts, % Day 15 Cells' quantity, 70.8 ± 6.0  36.8 ± 5.0*  66.1± 6.5*  10¹¹/L Non-differentiated 0.5 ± 0.1  0* 1.2 ± 0.2  blasts, %Proerythroblasts, % 0 0 0.1 ± 0.05 Erythroblasts, % 0.8 ± 0.2 0.4 ± 0.1 2.2 ± 0.5* Pronormoblasts, % 0.6 ± 0.1 0.3 ± 0.1  0.5 ± 0.1  Basophilic7.6 ± 0.8 2.4 ± 0.5* 6.8 ± 1.1  normoblasts, % Polychromatic 8.3 ± 1.13.3 ± 0.9* 15.7 ± 2.6*  normoblasts, % Eosinophilic 5.7 ± 1.1 3.2 ± 0.9*7.7 ± 2.2* normoblasts, % Red blood 0.58 ± 0.11 0.22 ± 0.12* 0.71 ±0.14* mitosis count, % Myeloblasts, % 2.0 ± 0.4 1.5 ± 0.4   4.0 ± 0.8**Promyelocytes, % 4.1 ± 0.6 1.5 ± 0.3* 5.0 ± 0.8* Myelocytes, % 5.9 ± 0.83.1 ± 0.6* 6.8 ± 0.7* Metamyelocytes, % 8.4 ± 1.2 3.3 ± 0.9   6.5 ±0.8** Stab neutrophils, % 11.5 ± 1.5  3.5 ± 0.6* 11.3 ± 1.3*  Segmented16.2 ± 2.2  42.1 ± 5.8  33.1 ± 4.7*  neutrophils, % Eosinophils, % 7.4 ±1.3 3.2 ± 0.8* 4.0 ± 1.5* Basophils, % 0 0 0 White blood 0.42 ± 0.100.17 ± 0.05* 0.57 ± 0.13* mitosis count, % Prolymphocytes, % 0 0 0.4 ±0.2* Lymphocytes, % 8.0 ± 0.6 2.0 ± 0.4  14.3 ± 1.2*  Plasma cells, %1.4 ± 0.3 0 2.6 ± 0.6* Promonocytes, % 0 0 0.1 Monocytes, % 0.8 ± 0.2 0.1 ± 0.05* 0.5 ± 0.2* Reticulocytes, % 2.4 ± 0.6 1.6 ± 0.4  4.1 ± 0.8*Megakaryocytes and 0.50 ± 0.10 0.15 ± 0.05* 0.54 ± 0.1* megakaryoblasts, %

[0633] TABLE 13 Effect of GSSG and Li•GSSG•Pt on cytokines' andhemopoietic factors' production by splenocytes, bone marrow, spleen andblood cellular indices, and bone marrow and spleen hematopoietic colonyformation capability in irradiated mice. (M ± m) Sham-irradiated animalsIrradiated animals Parameter N Normal saline Normal saline GSSGLi•GSSG•Pt Blood leukocyte 12 12.7 ± 1.3   3.4 ± 0.9*   6.7 ± 1.3◯  10.7± 2.0◯  count, 10⁹/L Blood lymphocyte 12  7.9 ± 0.7   2.2 ± 1.3*   5.2 ±0.8◯  7.4 ± 0.8◯  count, 10⁹/L Bone marrow 12 45.1 ± 3.2   14.0 ± 1.0*  23.3 ± 5.2◯  42.0 ± 4.0◯  nucleated cell number, 10⁶/LSplenocytes'count, 12  9.8 ± 1.5   4.8 ± 1.3*   6.3 ± 1.2◯  8.9 ± 2.0◯ 10⁷/organ Bone marrow CFU 12 59.4 ± 3.2   11.6 ± 2.2*   34.3 ± 3.9◯ 56.3 ± 3.9◯  Spleen CFU 12 93.2 ± 4.1   40.0 ± 5.4*   56.3 ± 6.8◯  89.6± 4.7◯  IL-1β 12   56 ± 3.2   8.3 ± 1.5*   1.8 ± 2.5◯  52.7 ± 5.4◯  IL-212   73 ± 6.2   10.6 ± 1.4*    28 ± 4.0   70.6 ± 7.1◯  IL-3 12  169 ±16.7  41.7 ± 4.7*   82.4 ± 7.9◯   167 ± 16.1◯ IL-4 12   86 ± 10.4 136.3± 12.9* 126.8 ± 6.4    84 ± 8.2◯  IL-6 12   98 ± 8.7   19.6 ± 1.8*  42.7 ± 4.1◯  95.7 ± 9.4◯  IL-8 12  103 ± 10.5  25.4 ± 2.4*   57.6 ±5.9   99.8 ± 9.9◯  IL-10 12   96 ± 10.3 154.8 ± 14.9* 132.9 ± 8.4◯  98.0± 9.9◯  IL-12 12   97 ± 8.7   28.7 ± 2.7*   48.3 ± 4.6   95.5 ± 9.5◯ INF-α 12  118 ± 11.4  29.4 ± 3.7*   56.2 ± 5.9◯ 105.6 ± 9.1◯  IFN-γ 12 116 ± 12.9  35.8 ± 3.2*   47.6 ± 4.3  113.8 ± 11.0◯ TNF-α 12   95 ±9.5   21.4 ± 2.6*   34.5 ± 3.8   91.7 ± 9.3◯  TNF-γ 12  115 ± 10.6  22.7± 2.5*   47.4 ± 5.1◯ 111.3 ± 12.3◯ GM-CSF 12  173 ± 17.2  39.2 ± 3.6* 127.5 ± 12.4 169.2 ± 16.7◯ G-CSF 12  156 ± 15.3  28.7 ± 2.7*    59 ±5.6◯ 139.4 ± 13.5◯ M-CSF 12  121 ± 12.6  45.2 ± 4.8*    78 ± 7.5◯ 118.7± 11.1◯

[0634] TABLE 14 Molecular mechanisms of the immunomodulating effects ofthe GSSG.Pt composite, indicating the reproduction of cytokines' effectsPhosphorilating level on tyrosine of the lymphocytes' The studyCytokines' content (g/ml) CD25+ content, cytosol proteins, conditionsIL-1 IL-6 TNF- IFN- (%) (impulse/min) 1 2 3 4 5 6 7 0 45 ± 4  120 ± 7 90± 7  90 ± 6 3.7 ± 0.9 6640 ± 270 (zero point as control) 10 50 ± 6  126± 9 89 ± 4 101 ± 9 3.6 ± 0.5 19240 ± 360* 30 48 ± 7  128 ± 6 93 ± 6 102± 8 3.8 ± 0.4 46980 ± 620* 1 hr. 52 ± 6   142 ± 11 108 ± 6  134 ± 6 3.9± 0.5 22350 ± 370* 6 hrs. 174 ± 17*  392 ± 12 402 ± 8*  214 ± 22* 4.1 ±0.4 11210 ± 260* 12 hrs. 275 ± 39*  2132 ± 132*  1525 ± 163*  514 ± 56* 5.9 ± 0.5* 8420 ± 170 24 hrs. 251 ± 23*  1621 ± 36* 1021 ± 56*  496 ±36* 17.6 ± 2.3* 6780 ± 420 48 hrs. 189 ± 17*  1241 ± 12*  893 ± 43*  247± 21 20.5 ± 4.3* 6320 ± 210

[0635] TABLE 15 Influence of the GSSG•Pt composite salts on thephosphorilating level on tyrosine and percentage of thelymphocytes-carriers of the IL-2-receptors to the total lymphocytes ofthe CBA mice in conditions of cyclophosphamide-induced immunodepression.Content of the Phosphorilating lymphocytes-carriers level on tyrosineThe tested articles of the IL-2-receptors (%) (impulse/min.) Na-GSSG•Pt16.8 ± 1.1* 28980 ± 420* Li-GSSG•Pt 20.2 ± 1.9* 34550 ± 790* Mg-GSSG•Pt18.6 ± 0.8* 29800 ± 880* Normal saline (control) 4.3 ± 1.3 11710 ± 340 

[0636] TABLE 16 Effect of the GSSG drug on blood and immunology indicesand cytokine levels in patient with stomach cancer, peritonealmetastases, ascites and splenomegaly. 2 months after the Parameter Priorto the treatment treatment beginning Hematology Erythrocytes, 10¹²/L 3.23.7 Hemoglobin, g/L 112 121 Platelets, 10⁹/L 205 195 Leukocytes, 10⁹/L12.4 8.9 Neutrophils (stab), % 12 8 Neutrophils (segm.), % 54 44Eosinophils, % 5 4 Lymphocytes, % 21 36 Monocytes, % 8 7 ESR, mm/hr 5415 Biochemistry Total protein, g/L 62 76 Albumin, % 26 42 α₁-globulin, %3 7 α₂-globulin, % 14 12 β-globulin, % 7 10 γ-globulin, % 50 26 A/Gratio 0.35 0.72 Urea, mmol/L 6.6 6.1 Creatinin, mmol/L 0.11 0.09Bilirubin, memol/L 40.0 32.4 Bilirubin conjugated, 31.0 21.4 μmol/LProthrombin index, % 75 79 Glucose, mmol/L 5.9 5.3 SGOT, mmol/hr/L 4.81.21 SGPT, mmol/hr/L 3.8 1.21 Immunology Lymphocytes, 10⁶/L 260.4 1204helpers (D4⁺), 10⁶/L 132.8 524 suppressors (D8), 10⁶/L 13 374(D4⁺)/(D8⁺) 10.2 1.4 NK-cells (CD16⁺), 10⁶/L 26 224 B-lymphocytes (D20⁺)26 152 IL2-receptor bearing cells 26.8 398 (D25⁺), 10⁶/L L 11-receptorbearing cells, 13 158 10⁶/L IgA, g/L 3.2 2.38 IgG, g/L 21.82 14.34 IgM,g/L 3.6 0.58 Cytokines' status IL-2, pg/mL 145 367 IL-1β, pg/mL 92 527IL-6, pg/mL 118 506 IFN-γ, pg/mL 105 624 TNF-α, pg/mL 183 507 GM-CSF,colonies/10⁵ cells 43.5 108

[0637] TABLE 17 Effect of the GSSG•Pt drug on blood and immunologyindices and cytokine levels in patient with stomach cancer, peritonealmetastases, ascites and splenomegaly. 2 months after the Parameter Priorto the treatment treatment beginning Hematology Erythrocytes, 10¹²/L 3.14.4 Hemoglobin, g/L 110 135 Platelets, 10⁹/L 215 275 Leukocytes, 10⁹/L12.2 8.1 Neutrophils (stab), % 11 2 Neutrophils (segm.), % 57 47Eosinophils, % 4 3 Lymphocytes, % 22 39 Monocytes, % 6 9 ESR, mm/hr 5415 Biochemistry Total protein, g/L 64 82 Albumin, % 21 50 α₁-globulin, %3 11 α₂-globulin, % 15 7 β-globulin, % 6 13 γ-globulin, % 50 19 A/Gratio 0.26 1.0 Urea, mmol/L 6.5 7.4 Creatinin, mmol/L 0.10 0.82Bilirubin, memol/L 36.0 20.1 Bilirubin conjugated, 28.7 14.4 μmol/LProthrombin index, % 73 95 Glucose, mmol/L 6.1 4.2 SGOT, mmol/hr/L 3.80.21 SGPT, mmol/hr/L 3.2 0.17 Immunology Lymphocytes, 10⁶/L 476.2 3320helpers (D4⁺), 10⁶/L 157.2 1454 suppressors (D8), 10⁶/L 15.1 908(D4⁺)/(D8⁺) 10.4 1.6 NK-cells (CD16⁺), 10⁶/L 39 776 B-lymphocytes(D20⁺)44 398 IL2-receptor bearing cells 42 2000 (D25⁺), 10⁶/L L 11-receptorbearing cells, 45 754 10⁶/L IgA, g/L 3.0 2.42 IgG, g/L 24.7 13.2 IgM,g/L 2.8 0.4 Cytokines' status IL-2, pg/mL 214 1237 IL-1β, pg/mL 115 1113IL-3, pg/mL 87 589 IL-4, pg/mL 230 108 IL-6, pg/mL 215 1553 IL-8, pg/mL136 157 IL-10, pg/mL 432 116 IL-12, pg/mL 89 626 IFN-α, pg/mL 86.4 962IFN-γ, pg/mL 129 919 TNF-α, pg/mL 202 1080 TNF-γ, pg/mL 163 745 GM-CSF,colonies/10⁵ cells 45.3 213 G-CSF, colonies/10⁵ cells 32.7 174 M-CSF,colonies/10⁵ cells 25.6 146

[0638] TABLE 18 GSSG•Pt influence on development of hematologic,biochemical, immunologic indices and cytokines' content of the patienthaving lung cancer Before Indexes treatment 3 months later HematologyErythrocytes (× 10¹²/L) 3.2 3.9 Hemoglobin, g/L 91 118 Leukocytes (×10⁹/L) 12.9 9.1 Lymphocytes, % 15 39 ESR, mm/hr 65 27 Biochemistry ALT(mmol/hr · L.) 0.7 0.59 AST (mmol/hr · L.) 1.8 0.5 Bilirubin - total(μm/L) 17.4 7.0 Urea, mmol/L 8.4 4.4 Creatinine, mmol/L 0.11 0.066Immunology helpers, (D4⁺), 10⁶/L 325 692 suppressors, (D8⁺), 10⁶/L 112320 NK-cells (CD16+), 10⁶/L 138 194 B-lymphocytes (D20⁺) 192 260 Cellsbearing IL-2 receptors (D25⁺), 141 236 10⁶/L Cytokine status IL-2, pg/mL159 360 IL-1β, pg/mL 120 375 IFN-γ, pg/mL 198 213 TNF-α pg/mL 535 822

[0639] TABLE 19 GSSG•Pt influence on development of hematologic,biochemical, immunologic indices and cytokines' content of the patienthaving lung cancer complicated with the liver metastases Indexes Beforetreatment 3 months later Hematology Erythrocytes (× 10¹²/L) 2.7 4.2Hemoglobin, g/L 62 141 Platelets, 10⁹/L 336 302 Leukocytes (× 10⁹/L)14.8 8.9 Stab neutrophils, % 20.5 2 Segmented neutrophils, % 42 59Lymphocytes, % 18.5 36 ESR, mm/hr 43 13 Biochemistry ALT (mmol/hr · L.)1.4 0.16 AST (mmol/hr · L.) 1.0 0.1 Bilirubin - total (μm/L) 24.0 6.2Urea, mmol/L 5.0 4.0 Creatinine, mmol/L 110 100 Immunology lymphocytes(CD3+), % 41 57 lymphocytes (CD3+), 10⁶/L 1148 1824 helpers (D4), % 1326 helpers (D4), 10⁶/L 364 532 suppressors (D8), % 14 16 suppressors(D8), 10⁶/L 292 412 D4⁺/D8⁺ 0.93 1.6 NK-cells (CD16+), 10⁶/L 98 454Cells bearing IL-2 receptors, (D25⁺), 120 460 10⁶/L B-lymphocytes(D72⁺), % 7 13 B-lymphocytes (D72⁺), 10⁶/L 196 416 Cytokine statusIL-1β, pg/mL 220 413 IL-2, pg/mL 150 492 IL-4, pg/mL 230 184 IL-6, pg/mL173 354 IL-10, pg/mL 918 626 IFN-α, pg/mL 284 383 IFN-γ, pg/mL 217 584TNFα 168 835

[0640] TABLE 20 Influence of GSSG•Pt on changes of hematologic,biochemical, serologic immune indices and cytokines' content at thepatient with chronic HBV Parameter Prior to the treatment 1 month afterthe treatment Hematology Erythrocytes 4.1 4.3 (× 10¹²/L) Hemoglobin, g/L100 115 Leukocytes 9.9 6.2 (× 10⁹/L) Lymphocytes, % 16 25 Stabneutrophils, % 8 3 Segmented 65 62 neutrophils, % Monocytes, % 9 7Eosinophils, % 2 3 ESR, mm/hr 35 17 Blood Biochemistry ALT (mmol/hr ·L.) 5.4 2.2 Bilirubin - total 34.6 26.0 (μm/L) Serology HBs Ag (ng/mL)183 178 Anti HBcor IgG +++ +++ Anti HBcor IgM − − PCR HBV + − Anti HBsAg <10 U/mL <10 U/mL Immunology D4⁺, 10⁶/L 325 692 D8⁺, 10⁶/L 112 320CD16⁺, 10⁶/L 138 94 D72⁺, 10⁶/L 192 160 95, (Fas Ag), % 10 21 Cytokinestatus IL-1β, pg/mL 187 97 IL-2, pg/mL 138 79 INF-γ, pg/mL 283 252

[0641] TABLE 21 Influence of GSSG•Pt on changes of hematologic,biochemical, serologic, immune indices and cytokines' content at thepatient with chronic HBV Parameter Prior to the treatment 1 month afterthe treatment Hematology Erythrocytes 3.8 4.5 (× 10¹²/L) Hemoglobin, g/L105 130 Leukocytes 10.5 5.6 (× 10⁹/L) Lymphocytes, % 19 28 Stabneutrophils, % 10 3 Segmented 42 64 neutrophils, % Monocytes, % 22 3Eosinophils, % 7 2 ESR, mm/hr 42 11 Blood Biochemistry ALT (mmol/hr ·L.) 6.2 0.8 Bilirubin - total 78.3 12.0 (μm/L) Serology HBs Ag (ng/mL)198 69 Anti HBcor IgG +++ +++ Anti HBcor IgM − − PCR HBV + − Anti HBs Ag<10 U/mL <10 U/mL Immunology D4⁺, 10⁶/ 306 785 D8⁺ 10⁶/ 121 528 CD16⁺,10⁶/ 143 85 D20⁺, 10⁶/ 260 144 95 + (FasAg)% 7 75 Cytokine status IL-1β,pg/mL 195 76 IL-2, pg/mL 156 59 IL-6, pg/mL 124 323 IL-4, pg/mL 1550 300IL-10, pg/mL 1362 686 INF-γ, pg/mL 275 192 TNF-α, pg/mL 720 184

[0642] TABLE 22 Changes in hematological, serological and biochemicalparameters before and after the treatment with the GSSG•Pt drugs use atthe patient with acute viral hepatitis B Prior to the After the 1 monthafter Parameter treatment treatment the treatment HematologyErythrocytes (× 10¹²/L) 3.9 4.1 4.2 Hemoglobin, g/L 116 125 134Leukocytes (× 10⁹/L) 4.5 4.7 4.6 Lymphocytes, % 46 38 35 Stabneutrophils, % 6 6 5 Segmented neutrophil, s % 42 51 58 Monocytes, % 3 21 Platelets, (thousand × 10⁹/L) 120 240 236 Eosinophils, % 3 3 1Serology HBs Ag (ng/mL) 129 117 − HBcor IgG +++ +++ +++ HBcor IgM + − −PCR HBV + + − Anti HBs Ag 10 U/ml 10 U/ml 10 U/ml PCR HDV + + − BloodBiochemistry Bilirubin (μm/L) 19.0 13.0 12.0 ALT (mmol/hr · L.) 2.8 0.090.38

[0643] TABLE 23 Patient's immunologic status at the treatment with theGSSG•Pt drugs at the patient with acute viral hepatitis B Index Beforethe treatment After the treatment CD4⁺ 680 504.5 CD8⁺ 467 560.7CD4⁺/CD8⁺ 1. 0.93 CD4⁺CD8⁺ 363 256 CD16⁺ 595.7 378.4 CD72⁺ 483.4 485.9CIC 112.85 93.2 HLADR 513 467 FasAg (CD95+), % 1.3 23

[0644] TABLE 24 Patient's cytokine status at the treatment with thcGSSG•Pt drugs at the patient with acute viral hepatitis B Before thetreatment After the treatment Index (pg/mL) (pg/mL) IL-1β 296.5 98.5IL-2 121 92 IL-6 189 260 IL-10 1001 226 IFN-γ 350.9 108 IL-4 1650 450TNF-α 1073 158

[0645] TABLE 25 Changes in hematological, scrological and biochemicalparameters before and after the treatment with the GSSG•Pt drug use atthe patient with chronic viral hepatitis C Prior to 1 month 3 months theAfter the after the after the Parameter treatment treatment treatmenttreatment Hematology Erythrocytcs 4.1 4.0 4.4 4.6 (× 10¹²/L) Hemoglobin,g/L 120 140 136 148 Leukocytes 8.9 6.7 5.7 5.8 (× 10⁹/L) Lymphocytes, %30 47 46 17 Stab neutrophils, % 5 2 1 2 Segmented 52 41 38 68neutrophils, % Monocytes, % 10 8 11 12 Platelets 240 280 215 265(thousand × 10⁹/L) Eosinophils, % 3 2 4 1 Serology HBs Ag (ng/mL) − − −− Anti HBcor IgG +++ +++ +++ +++ Anti HBcor IgM − − − − PCR HCV + − − −Anti HBs Ag 10 U/ml 10 U/ml 75 U/ml 75 U/ml Anti HCV IgG +++cor +++cor+++cor++ns +++cor++ns Blood Chemistry Bilirubin (μmol/L) 34.0 22.0 24.018.0 ALT (mmol/hr · L.) 1.8 0.52 0.18 0.3

[0646] TABLE 26 Patient's immunologic status before and after thetreatment with the GSSG•Pt drugs at the patient with chronic viralhepatitis C Index Before the treatment After the treatment CD4⁺ 519 679CD8⁺ 541 450 CD4⁺/CD8⁺ 1 1.23 CD4⁺CD8⁺ 363 2568 CD16⁺ 573.7 358 CD72⁺676 459 CIC 180,7 92 HLA-DR 715 424 CD95⁺ (FasAg), % 5 45

[0647] TABLE 27 Patient's cytokine status at the treatment with theGSSG•Pt drugs at the patient with chronic viral hepatitis C Before thetreatment After the treatment Index (pg/mL) (pg/mL) IL-1β 239,5 108,3IL-2 128 88 IL-6 156 250 IL-10 1133 887 IFN-γ 307,9 280 IL-4 1800 600TNF-α 976 358

[0648] TABLE 28 Results of the GSSG and GSSG•Pt action on normallymphocytes' count in vitro during 48-hour incubation* (M ± m) (<0.05).Tested articles 24 hours, 48 hours, (concentration 100 μg/mL) Cells'state 10³ 10³ Control Cells - total 264 ± 30  285 ± 36 Dead cells - %7.0 ± 1.2 12.0 ± 1.4 GSSG Cells - total 265 ± 34  280 ± 38 Dead cells -% 8.0 ± 1.5 13.0 ± 1.3 GSSG•Pt Cells - total 269 ± 32  287 ± 35 Deadcells - % 6.0 ± 1.1 12.0 ± 1.6

[0649] TABLE 29 HL-60 cells' growth development during 48-hourincubation after the GSSG and GSSG•Pt treatment* (M ± m) (<0.05). Testedarticles (concentration 100 24 hours, 48 hours, μg/mL) Cells' state 10³10³ Control Cells - total 800 ± 30   2785 ± 36.1  Dead cells - % 3.0 ±1.2 6.0 ± 1.4 GSSG Cells - total 515 ± 54  780 ± 38  Dead cells - % 27.0 ± 3.3**   53 ± 6.3** GSSG•Pt Cells - total 360 ± 32  283 ± 35 Dead cells - %  87.0 ± 8.5** 100**

[0650] TABLE 30 Results of the GSSG•Pt action on normal lymphocytes'count in vitro during 48-hour incubation* (M ± m) (<0.05). GSSC•Pt, 24hours, 48 hours, μg/mL Cells' state 10³ 10³ Control Cells - total 264 ±30  285 ± 36 Dead cells - % 7.0 ± 1.2 12.0 ± 1.4 DNA apoptoticdegradation − − 10 Cells - total 266 ± 28  285 ± 34 Dead cells - % 7.5 ±1.5 11.0 ± 2.0 DNA apoptotic degradation − − 100 Cells - total 269 ± 32 287 ± 35 Dead cells - % 6.0 ± 1.1 12.0 ± 1.6 DNA apoptotic degradation −−

[0651] TABLE 31 HL-60 cells' growth development during 48-hourincubation after the GSSG•Pt treatment* (M ± m) (<0.05). GSSG•Pt(concentration, 24 hours, 48 hours, μg/mL) Cells' state 10³ 10³ ControlCells - total 800 ± 30   2785 ± 36.1  Dead cells - % 3.0 ± 1.2 6.0 ± 1.4DNA apoptotic − −+ degradation 10 Cells - total 611 ± 52  778 ± 35  Deadcells - %  26.0 ± 4.2**   49 ± 5.0** DNA apoptotic + + degradation 100Cells - total 360 ± 32  283 ± 35  Dead cells - %  87.0 ± 8.5** 100** DNAapoptotic + + degradation

[0652] TABLE 32 C-8 cells' growth development during 48-hour incubationafter the GSSG•Pt treatment* (M ± m) (<0.05). GSSG•Pt (concentration, 24hours, 48 hours, μg/mL) Cells' state 10³ 10³ Control Cells - total 139.0± 14   208.0 ± 24   Dead cells - %  1.5 ± 0.5 6.0 ± 1.5 DNA apoptotic −− degradation 10 Cells - total 54.4 ± 3.8 59.7 ± 4.6  Dead cells - % 32.0 ± 4.0**   52 ± 6.0** DNA apoptotic + + degradation 100 Cells -total 32.6 ± 3.9 22.1 ± 2.8  Dead cells - %  76.3 ± 7.8** 100** DNAapoptotic + + degradation

[0653] TABLE 33 A-4 cells' growth development during 48-hour incubationafter the GSSG•Pt treatment* (M ± m) (<0.05). GSSG•Pt (concentration, 24hours, 48 hours, μg/mL) Cells' state 10³ 10³ Control Cells - total 98.1± 7.2  197.0 ± 18.2  Dead cells - % 2.0 ± 0.5 4.5 ± 1.5 DNA apoptotic −− degradation 10 Cells - total 53.7 ± 4.1  61.4 ± 7.5  Dead cells - % 33.0 ± 3.0**   55 ± 5.0** DNA apoptotic + + degradation 100 Cells -total 24.7 ± 2.5  13.7 ± 2.4  Dead cells - %  61.6 ± 6.5** 100** DNAapoptotic + + degradation

[0654] TABLE 34 C-8 cells' growth development during 48-hour incubationafter the GSSG•Pt treatment* (M ± m) (<0.05). GSSG•Pt (concentration, 24hours, 48 hours, μg/mL) Cells' state 10³ 10³ Control Cells - total 121.0± 11.0  217.5 ± 15.3  Dead cells - % 1.5 ± 0.5 6.0 ± 1.5 DNA apoptotic −− degradation 100 Cells - total 32.6 ± 3.3  22.1 ± 2.8  Dead cells - % 76.3 ± 7.8** 100** DNA apoptotic + + degradation

[0655] TABLE 35 Growth development of the p21-knockout (p21-) cellsduring 48-hour incubation after the GSSG•Pt treatment* (M ± m) (<0.05).GSSG•Pt (concentration, 24 hours, 48 hours, μg/mL) Cells' state 10³ 10³Control Cells - total 109.0 ± 9.6  191.0 ± 12.4  Dead cells - % 0.5 ±0.5 1.5 ± 0.5 DNA apoptotic − − degradation 100 Cells - total 46.7 ±5.3  31.2 ± 2.5  Dead cells - %  64.5 ± 6.5**  78.0 ± 7.3** DNAapoptotic + + degradation

[0656] TABLE 36 Blood glucose and biochemical blood indices highlycorrelated with it (r₁ and r₂ > 0.85)* in the patient with diabetesmellitus Before 1 month after 4 months after Indices treatment thetreatment the treatment Blood glucose, mmol/L 11.5-19.1 8.2-10.6 4.8-7.6cAMP/cGMP 7.8 6.5 4.1 TDR (thiol-disulfide ratio) 1.6 1.3 0.9

[0657] TABLE 37 Hematologic, biochemical and immunologic indices in thepatient with diabetes mellitus Prior to the 1 month after 4 months afterIndex treatment the treatment the treatment Hematology Erythrocytes,10¹²/L 3.9 4.1 4.4 Hemoglobin, g/L 120 131 143 Platelets, 10⁹/L 199 211263 Leukocytes, 10⁹/L 8.1 7.3 6.1 Stab netrophils, % 5 4 4 Segmentedneutrophils, 38 53 57 % Lymphocytes, 10⁹/L 52 37 34 Monocytes, % 3 3 4Eosinophils, % 2 2 1 ESR, mm/hour 17 13 10 Blood biochemistry ALT,/. . .0.49 0.37 0.28 AST,/. . . 0.32 0.36 0.31 Total protein g/L 75 71 72Total bilirubin, μmol/L 11.2 9.4 8.1 Total cholesterol, μmol/L 8.60 7.526.1 Triglycerides, μmol/L 4.8 3.9 2.7 Urea, mmol/l 4.7 4.3 3.8Creatinine, mmol/l 0.146 0.104 0.097 Immunity B-lymphocytes 486 409 391(CD20+), 10⁶/L T-helpers 1432 1109 932 (CD4+), 10⁶/L T-suppressors 1104969 710 (CD8+), 10⁶/L CD25⁺, 10⁶/L 463 537 499 Circulating immune 221156 102 complexes, U

What is claimed:
 1. A composite comprising an oxidized glutathione-basedcompound and a metal material in a molar equivalent ratio of betweenabout 3000:1 to about 1:1, wherein the metal material comprises a metalselected from the group consisting of platinum and palladium.
 2. Thecomposite of claim 1, wherein the composite comprises the oxidizedglutathione-based compound and the metal material in a molar equivalentratio of between about 1000:1 to about 1:1.
 3. The composite of claim 2,wherein the composite comprises the oxidized glutathione-based compoundand the metal material in a molar equivalent ratio of between about1000:1 to about 10:1.
 4. The composite of claim 2, wherein the compositecomprises the oxidized glutathione-based compound and the metal materialin a molar equivalent ratio of between about 1000:1 to about 100:1. 5.The composite of claim 1, wherein the metal is platinum.
 6. Thecomposite of claim 3, wherein the platinum material is selected from thegroup consisting of a platinum salt, a coordination compound and anorganometallic compound.
 7. The composite of claim 6, wherein theplatinum material is a platinum coordination compound.
 8. The compositeof claim 7, wherein the coordination compound is cis-platin.
 9. Thecomposite of claim 1, wherein the oxidized-based glutathione compoundhas the formula:

and salts of said formula; wherein A, B, D, E, G and H can be the sameor different and each is selected from the group consisting of anorganic unit and salts of the organic unit.
 10. The composite of claim9, wherein A, B, D, E, G and H can be the same or different and eachincludes a unit selected from the group consisting of amine groups,carboxyl groups and amides.
 11. The composite of claim 10, wherein anytwo of A, B, D, E, G and H are linked to each other by at least onecovalent bond.
 12. The composite of claim 11, wherein any two of A, B,D, E, G and H are linked to each other by an amide bond.
 13. Thecomposite of claim 10, wherein A, B, D, E, G and H can be the same ordifferent and each includes an amino acid.
 14. The composite of claim10, wherein the oxidized glutathione-based compound is oxidizedglutathione and both A and E are —CO₂H, both B and D are —NH₂ and both Gand H are —CO₂M, M being a counterion.
 15. The composite of claim 10,wherein the oxidized glutathione-based compound isS-thioethylamine•glutathione disulfide.
 16. The composite of claim 10,wherein the oxidized glutathione-based compound is bis-(DL-6,8-thioeticacid)•glutathione disulfide.
 17. The composite of claim 10, wherein theoxidized glutathione-based compound is (β-alanyl-L-histidyl)•glutathionedisulfide.
 18. The composite of claim 10, wherein the oxidizedglutathione-based compound is (9-β-D-ribofuranosyladenyl)•glutathionedisulfide.
 19. The composite of claim 10, wherein the oxidizedglutathione-based compound is bis-(L-2-amino-4-(methylthio)butanoicacid)•glutathione disulfide.
 20. The composite of claim 10, wherein theoxidized glutathione-based compound is bis-(L-phenylalanyl)•glutathionedisulfide.
 21. The composite of claim 10, wherein the oxidizedglutathione-based compound has an acylated primary glutamic acid aminogroup of oxidized glutathione.
 22. The composite of claim 21, whereinthe oxidized glutathione-based compound is selected from the groupconsisting of bis-(methionyl)•glutathione disulfide,bis-(aspartyl)•glutathione disulfide, bis-(histidyl)•glutathionedisulfide, bis-(3-iodine-tyrosyl)•glutathione disulfide,(γ-aminobutanoyl)•glutathione disulfide,bis-(γ-hydroxybutanoyl)•glutathione disulfide, bis-(lipoyl)•glutathionedisulfide, and bis-(3,4-dihydroxyphenylalaninyl)•glutathione disulfide.23. The composite of claim 8, wherein the oxidized glutathione-basedcompound has an amide or phosphoramide bond to a unit selected from thegroup consisting of heterocyclic carbonic acids and nucleotides.
 24. Thecomposite of claim 23, wherein the oxidized glutathione-based compoundis selected from the group consisting ofbis-(pyridine-3-carbonyl).glutathione disulfide,uridine-5′-monophosphatoyl•glutathione disulfide,inosine-5′-monophosphatoyl•glutathione disulfide,folliculylsuccinyl•glutathione disulfide andglycerol-1,3-diphosphatyl•glutathione disulfide.
 25. The composite ofclaim 10, wherein the oxidized glutathione-based compound is selectedfrom the group consisting of tetra-dopamine•glutathione disulfide andtheophylline•glutathione disulfide.
 26. The composite of claim 1,wherein the oxidized glutathione-based compound is chemically interactedwith the material comprising platinum.
 27. A method for stabilizing adisulfide bond of an oxidized glutathione-based compound, comprisinginteracting the oxidized glutathione-based compound with a materialcomprising platinum.
 28. The method of claim 27, wherein themetal-stabilized oxidized glutathione-based compound is GSSG.Pt.
 29. Themethod of claim 27, wherein the metal-stabilized oxidizedglutathione-based compound is a salt of GSSG.Pt.
 30. The method of claim27, wherein the platinum present in an amount of between about 0.0003molar equivalent to about 1 molar equivalent relative to the oxidizedglutathione-based compound.
 31. The method of claim 27, wherein theplatinum is present in an amount of between about 0.001 molar equivalentto about 0.01 molar equivalent relative to the oxidizedglutathione-based compound.
 32. The method of claim 27, wherein theplatinum is present in an amount of between about 0.001 molar equivalentto about 0.1 molar equivalent relative to the oxidized glutathione-basedcompound.
 33. The method of claim 27, wherein the platinum is present inan amount of between about 0.001 molar equivalent to about 1 molarequivalent relative to the oxidized glutathione-based compound.
 34. Themethod of claim 27, wherein the platinum is selected from the groupconsisting of platinum metal, a salt, a coordination compound and anorganometallic compound.
 35. The method of claim 34, wherein thematerial is cis-platin.
 36. The method of claim 27, wherein the oxidizedglutathione-based compound comprises the formula:

and salts of said formula; wherein A, B, D, E, G and H4 can be the sameor different and each is selected from the group consisting of anorganic unit and salts of the organic unit.
 37. The method of claim 27,wherein the interacting comprises: providing a glutathione-basedcompound; and reacting the glutathione-based compound with an oxidantand a material comprising platinum to form an oxidized glutathione-basedcompound having a stabilized disulfide bond.
 38. The method of claim 37,wherein the oxidant is selected from the group consisting of oxygen andhydrogen peroxide.
 39. The method of claim 38, wherein the oxidant ishydrogen peroxide.
 40. The method of claim 39, wherein the reacting stepcomprises reacting one molar equivalent of the glutathione-basedcompound with less than about 1 molar equivalent of the hydrogenperoxide and between about 0.0003 molar equivalent and about 1 molarequivalent of the material comprising platinum.
 41. The method of claim39, wherein the reacting step comprises reacting one molar equivalent ofthe glutathione-based compound with less than about 1 molar equivalentof the hydrogen peroxide and between about 0.001 molar equivalent andabout 0.1 molar equivalent of the material comprising platinum.
 42. Themethod of claim 39, wherein the reacting step comprises reacting onemolar equivalent of the glutathione-based compound with less than about1 molar equivalent of the hydrogen peroxide and between about 0.001molar equivalent and about 0.01 molar equivalent of the materialcomprising platinum.
 43. The method of claim 40, wherein the reactingstep comprises reacting one molar equivalent of the glutathione-basedcompound with less than about 1 molar equivalent of the hydrogenperoxide and between about 0.001 molar equivalent and about 1 molarequivalent of the material comprising platinum.
 44. The method of claim4 0, wherein the reacting step comprises reacting one molar equivalentof the glutathione-based compound with about 0.9 molar equivalent of thehydrogen peroxide and between about 0.0003 molar equivalent and about 1molar equivalent of the material comprising platinum.
 45. The method ofclaim 40, wherein the reacting step comprises reacting one molarequivalent of the glutathione-based compound with about 0.9 molarequivalent of the hydrogen peroxide and between about 0.001 molarequivalent and about 0.1 molar equivalent of the material comprisingplatinum.
 46. The method of claim 40, wherein the reacting stepcomprises reacting one molar equivalent of the glutathione-basedcompound with about 0.9 molar equivalent of the hydrogen peroxide andbetween about 0.001 molar equivalent and about 0.01 molar equivalent ofthe material comprising platinum.
 47. The method of claim 44, whereinthe reacting step comprises reacting one molar equivalent of theglutathione-based compound with about 0.9 molar equivalent of thehydrogen peroxide and between about 0.001 molar equivalent and about 1molar equivalent of the material comprising platinum.
 48. The method ofclaim 27, wherein the interacting comprises adding between about 0.0003molar equivalent to about 1 molar equivalent of the material comprisingplatinum to 1 about molar equivalent of the oxidized glutathione-basedcompound.
 49. The method of claim 27, wherein the oxidizedglutathione-based compound is a salt selected from the group consistingof alkali metal salts, alkaline earth metal salts and transition metalsalts.
 50. The method of claim 4 9, wherein the oxidizedglutathione-based compound is a salt selected from the group consistingof lithium salts, sodium salts, potassium salts, magnesium salts,calcium salts, vanadium salts, manganese salts, iron salts, molybdenumsalts and zinc salts.
 51. The method of claim 27, wherein the oxidizedglutathione-based compound is a fluoride-containing salt.
 52. A methodof stimulating endogenous production of cytokines and hemopoieticfactors comprising introducing to a mammalian body in need ofstimulation of cytokines or hemopoietic factors or both, an effectiveamount of a composite comprising an oxidized glutathione-based compoundand a metal material in a molar equivalent ratio of between about 3000:1to about 1:1 wherein the metal material comprises a metal selected fromthe group consisting of platinum and palladium, for a period of time tostimulate said endogenous production to obtain a therapeutic effect fora disease.
 53. The method of claim 52, wherein the molar equivalentratio is between about 1000:1 to about 1:1.
 54. The method of claim 52,wherein the molar equivalent ratio is between about 1000:1 to about10:1.
 55. The method of claim 52, wherein the molar equivalent ratio isbetween about 1000:1 to about 100:1.
 56. The method of claim 52, whereinthe metal is platinum.
 57. The method of claim 56, wherein the materialis cis-platin.
 58. The method of claim 52, wherein the composite isadministered orally.
 59. The method of claim 52, wherein the disease isselected from the group consisting of oncological, infectious,immunological, ischemic, neurodegenerative metabolic, endocrinal andother diseases.
 60. The method of claim 59, wherein the oncologicaldisease is selected from the group consisting of lung cancer, melanoma,cerebral tumors, colorectal cancer, breast cancer, prostate cancer,ovarian cancer, acute lymphoblastic leukosis and acute myeloblasticleukosis.
 61. The method of claim 59, wherein the infectious disease isselected from the group consisting of tuberculosis, viral hepatitis B,viral hepatitis C, mixed infections (HBV and HCV), herpes, meningitis(sepsis), peritonitis, acute pancreatis and supporative post-surgerysequalae.
 62. The method of claim 59, wherein the immunological diseaseis selected from the group consisting of AIDS, immunosuppressions ofinfectious origin, immunosuppressions of radiation origin,immunosuppressions of toxic origin, glomerulonephritis, rheumatoidarthritis, collagenosis, systemic lupus erythematosis and atopic formsof allergic conditions.
 63. The method of claim 59, wherein the ischemicdisease is selected from the group consisting of ischemic cerebralconditions and ischemic heart disease.
 64. The method of claim 59,wherein the neurodegenerative disease is selected from the groupconsisting of Alzheimer's disease, hereditary (Huntington's) chorea,amyotrophic lateral sclerosis, neuro-AIDS and demyelinating diseases.65. The method of claim 59, wherein the neurodegenerative disease is aneurobehavioral disease selected from the group consisting of narcoticabstinence, cerebral hypoxia, manic-depressive psychosis andschizophrenia.
 66. The method of claim 59, wherein the metabolic diseaseis atherosclerosis.
 67. The method of claim 59, wherein the endocrinaldisease is associated with hypothalamic-hypophysil-ovarian function. 68.The method of claim 52, wherein the therapeutic effect comprises aprocess selected from the group consisting of regulating proliferationin normal cells, regulating differentiation in normal cells and inducingapoptosis of transformed cells.
 69. The method of claim 52, wherein thecomposite is administered in a dosage of between about 0.1 mg/kg toabout 1.0 mg/kg by body weight.
 70. The method of claim 52, wherein thecomposite is administered in a dosage of between about 1 mg/m² to about100 mg/m² by body surface.
 71. The method of claim 52, wherein thecomposite is administered as a solution form selected from the groupconsisting of inhalation solutions, local instillations, eye drops,intranasal introductions, ointment for epicutaneous applications,intravenous solutions, injection solutions and suppositories.
 72. Themethod of claim 71, wherein the solution has a concentration of betweenabout 1% to about 10% of the composite.
 73. The method of claim 52,wherein the composite is administered as an injectable form.
 74. Themethod of claim 73, wherein the injectable form comprises the compositein a solution in a concentration of between about 0.01% to about 3.0%.75. The method of claim 72, wherein the composite is administered in adosage of between about 0.01 mg/kg to about 1.0 mg/kg by body weight.76. The method of claim 72, wherein the composite is administered in adosage of between about 1 mg/m² to about 100 mg/m² by body surface. 77.A method of enhancing and prolonging the ability of an oxidizedglutathione-based compound to stimulate endogenous production ofcytokine and hemopoietic factors, said method comprising interacting theoxidized glutathione-based compound with a metal material in a molarequivalent ratio of between about 3000:1 to about 1:1 wherein the metalmaterial comprises a metal selected from the group consisting ofplatinum and palladium.
 78. The method of claim 68, wherein said cellsare in a mammalian body and said composite is introduced into said bodyat a rate of from about 0.01 mg/kg to about 1.0 mg/kg of body weight atleast one time a day for at least one day.
 79. The method of claim 68,wherein said cells are in a mammalian body and said composite isintroduced topically to a topical area at a dose of from about 1.0 mg/m²to about 100 mg/m² of topical area.
 80. A method for treating a subjecthaving a disease, comprising: administering to the subject in need ofsuch treatment a composite comprising an oxidized glutathione-basedcompound and a metal material in a molar equivalent ratio of betweenabout 3000:1 to about 1:1 in an amount effective to stimulate endogenousproduction of cytokines and or hemopoietic factors or both, to obtain atherapeutic effect, wherein the metal material comprises a metalselected from the group consisting of platinum and palladium.
 81. Themethod of claim 80, wherein the molar equivalent ratio is between about1000:1 to about 1:1.
 82. The method of claim 80, wherein the molarequivalent ratio is between about 1000:1 to about 10:1.
 83. The methodof claim 80, wherein the molar equivalent ratio is between about 1000:1to about 100:1.
 84. The method of claim 80, wherein the disease is lungcancer and the composite is GSSG.Pt.
 85. The method of claim 80, whereinthe disease is melanoma and the composite isbis-(-iodine-tyrosyl)-GSSG.Pt.
 86. The method of claim 80, wherein thedisease is a cerebral tumor and the composite is bis-(dopamine)-GSSG.Pt.87. The method of claim 80, wherein the disease is a colorectal cancerand the composite is bis-(cysteamine)-GSSG.Pt.
 88. The method of claim80, wherein the disease is breast cancer and the composite iscysteamine-GSSG.Pt.
 89. The method of claim 80, wherein the disease isprostate cancer and the composite is dizinc salts of GSSG.Pt.
 90. Themethod of claim 80, wherein the disease is ovarian cancer and thecomposite is theophylline-GSSG.Pt.
 91. The method of claim 80, whereinthe disease is acute lymphoblastic leukosis and the composite is alithium salt of GSSG.Pt.
 92. The method of claim 80, wherein the diseaseis acute myeloblastic leukosis and the composite is selected from thegroup consisting of dilithium salt of GSSG.Pt and cysteamine-GSSG.Pt andcombinations thereof.
 93. The method of claim 80, wherein the disease istuberculosis and the composite is bis-(histidyl)-GSSG.Pt.
 94. The methodof claim 80, wherein the disease is selected from the group consistingof viral hepatitis B, viral hepatitis C, and mixed-infections thereofand the composite is selected from the group consisting of GSSG.Pt andinosine-5-monophosphatyl-GSSG.Pt.
 95. The method of claim 80, whereinthe disease is herpes and the composite is selected from the groupconsisting of GSSG.Pt and inosine-5-monophosphatyl-GSSG.Pt.
 96. Themethod of claim 80, wherein the disease is meningitis, sepsis and thecomposite is tetra-dopamine-GSSG.Pt.
 97. The method of claim 80, whereinthe disease is peritonitis and the composite is selected from the groupconsisting of GSSG.Pt and tetra-dopamine-GSSG.Pt and combinationsthereof.
 98. The method of claim 80, wherein the disease is acutepancreatitis and the composite is selected from the group consisting ofGSSG.Pt and inosine-5-monophosphatyl-GSSG.Pt and combinations thereof.99. The method of claim 80, wherein the disease is suppurativepost-surgery sequalae and the composite is selected from the groupconsisting of GSSG.Pt and inosine-5-monophosphatyl-GSSG.Pt andcombinations thereof.
 100. The method of claim 80, wherein the diseaseis AIDS and the composite is selected from the group consisting ofGSSG.Pt and uridine-(5-monophosphatyl)-GSSG.Pt and combinations thereof.101. The method of claim 80, wherein the disease is immunosuppressionsof infectious origin and the composite is selected from the groupconsisting of GSSG.Pt and uridine-(5-monophosphatyl)-GSSG.Pt andcombinations thereof.
 102. The method of claim 80, wherein the diseaseis glomerulonephritis and the composite is selected from the groupconsisting of GSSG.Pt and a lithium salt of GSSG.Pt and combinationsthereof.
 103. The method of claim 80, wherein the disease is selectedfrom the group consisting of rheumatoid arthritis and the composite isselected from the group consisting of GSSG.Pt and a lithium salt ofGSSG.Pt and combinations thereof.
 104. The method of claim 80, whereinthe disease is collagenosis and the composite is selected from the groupconsisting of GSSG.Pt and a lithium salt of GSSG.Pt and combinationsthereof.
 105. The method of claim 80, wherein the disease is systemiclupus erythematosus and the composite is selected from the groupconsisting of GSSG.Pt and a lithium salt of GSSG.Pt and combinationsthereof.
 106. The method of claim 80, wherein the disease is an atopicform of an allergic condition and the composite is selected from thegroup consisting of GSSG.Pt and dihydrofluoride-GSSG.Pt and combinationsthereof.
 107. The method of claim 80, wherein the disease isdiabetes-type I and the composite is a vanadium salt of GSSG.Pt. 108.The method of claim 80, wherein the disease is diabetes-type II and thecomposite is bis-(lipoyl)-GSSG.Pt.
 109. The method of claim 80, whereinthe disease is an ischemic cerebral condition and the composite isbis-(phenylalanyl)-GSSG.Pt.
 110. The method of claim 80, wherein thedisease is an ischemic heart disease and the composite isbis-(carnosyl)-GSSG.Pt.
 111. The method of claim 80, wherein the diseaseis an ischemic heart disease manifested mainly as a syndrome offunctional myocardial failure and the composite isglycerol-(1,3-diphosphatyl)-GSSG.Pt.
 112. The method of claim 80,wherein the disease is neurodegenerative disease and the composite isbis-(3,4-dihydroxyphenylalanyl)-GSSG.Pt.
 113. The method of claim 80,wherein the disease is demyelinating disease and the composite isbis-(3,4-dihydroxyphenylalanyl)-GSSG.Pt.
 114. The method of claim 80,wherein the disease is cerebral hypoxia and the composite isgamma-hydroxy-(butanoyl)-GSSG.Pt.
 115. The method of claim 80, whereinthe disease is manic-depressive psychosis and the composite isgamma-amino-(butanoyl)-GSSG.Pt.
 116. The method of claim 80, wherein thedisease is metabolic disease and the composite isbis-(nicotinoyl)-GSSG.Pt.
 117. The method of claim 8 0, wherein thedisease is an endocrinal disease and the composite isfolliculyl-(succinyl)-GSSG.Pt.
 118. The method of claim 120, whereinsaid normal and transformed cells are in a mammalian body and saidcomposite is introduced into said body at a rate of from about 0.01mg/kg to about 1.0 mg/kg of body weight at least one time a day for atleast one day.
 119. The method of claim 120, wherein said normal andtransformed cells are in a mammalian body and said composite isintroduced topically to a topical area at a dose of from about 1.0 mg/m²to about 100 mg/m² of topical area.
 120. The method of claim 77, whereinthe enhancement and prolonging of the ability of the oxidizedglutathione-based compound to stimulate endogenous production ofcytokine and hemopoietic factors comprises a process selected from thegroup consisting of regulating proliferation in normal cells, regulatingdifferentiation in normal cells and inducing apoptosis of transformedcells.
 121. The composite of claim 1, wherein the composite is presentin a dosage form for therapeutic use.
 122. The composite of claim 121,wherein the dosage is between about 0.10 mg/kg to about 1.0 mg/kg bybody weight.
 123. The composite of claim 121, wherein the dosage isbetween about 1 mg/m² to about 100 mg/m² by body surface.
 124. Thecomposite of claim 121, wherein the composite is capable of beingintroduced topically and the dosage is between about 1 mg/m² to about100 mg/m² of topical area.
 125. The composite of claim 122, wherein thecomposite is capable of being introduced in injectable form in aconcentration between about 1% to about 10% by weight/volume.
 126. Thecomposite of claim 122, wherein the composite is capable of beingintroduced in injectable form in a concentration between about 0.01% toabout 3.0% by weight/volume.
 127. The composite of claim 1, wherein thecomposite is GSSG.Pt.
 128. The composite of claim 1, wherein thecomposite is a salt of GSSG.Pt.
 129. The method of claim 52, wherein thecomposite is GSSG.Pt.
 130. The method of claim 52, wherein the compositeis a salt of GSSG.Pt.
 131. The method of claim 77, wherein the compositeis GSSG.Pt.
 132. The method of claim 77, wherein the composite is a saltof GSSG.Pt.
 133. The method of claim 80, wherein the composite isGSSG.Pt.
 134. The method of claim 80, wherein the composite is a salt ofGSSG.Pt.
 135. The composite of claim 123, wherein the composite iscapable of being introduced in injectable form in a concentrationbetween about 1% to about 10% by weight/volume.
 136. The composite ofclaim 123, wherein the composite is capable of being introduced ininjectable form in a concentration between about 0.01% to about 3.0% byweight/volume.
 137. A composite comprising an oxidized glutathione-basedcompound comprising a carbon/nitrogen backbone of each of a dimer of aglutamic acid bonded to a cysteine bonded to a glycine, the dimer beinglinked through a disulfide unit, said composite further comprising ametal material in a molar equivalent ratio of between about 3000:1 toabout 1:1, wherein the metal material comprises a metal selected fromthe group consisting of platinum and palladium.